Derivatives of amphotericin b

ABSTRACT

Disclosed are derivatives of amphotericin B (AmB) characterized by improved therapeutic index compared to AmB. The AmB derivatives include C16 ureas, carbamates, and amides according to Formula (I); C3′-substituted C16 ureas, carbamates, and amides according to Formula (II); C16 acyls according to Formula (III); C2′epi-C16 ureas, carbamates, and amides according to Formula (IV); and C16 oxazolidinone derivatives according to Formula (V). Also disclosed are pharmaceutical compositions comprising the AmB derivatives, and therapeutic methods of using the AmB derivatives.

RELATED APPLICATION

This application claims benefit of priority from U.S. Provisional PatentApplication No. 62/147,949, filed Apr. 15, 2015.

BACKGROUND OF THE INVENTION

For more than half a century amphotericin B (AmB) has served as the goldstandard for treating systemic fungal infections. AmB has a broadspectrum of activity, is fungicidal, and is effective even againstfungal strains that are resistant to multiple other agents.Surprisingly, clinically significant microbial resistance has remainedexceptionally rare while resistance to next generation antifungals hasappeared within just a few years of their clinical introduction.Unfortunately, AmB is also highly toxic. Deray, G, J AntimicrobChemother 49 Suppl 1: 37-41 (2002). Thus, the effective treatment ofsystemic fungal infections with AmB is all too often precluded, not by alack of efficacy, but by dose-limiting side effects. Mora-Duarte, J etal., N Engl J Med 347: 2020-9 (2002). Some progress has been made usingliposome delivery systems, but these treatments are prohibitivelyexpensive and significant toxicities remain. Wong-Beringer, A et al.,Clin Infect Dis 27: 603-18 (1998). Thus, a need exists for an effectivebut less toxic form or derivative of AmB.

SUMMARY OF THE INVENTION

An aspect of the invention is a compound represented by Formula (I) or apharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X is —N(R²)—, —C(R³)(R³)—, or —O—;

R² is hydrogen or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, aminoalkyl, and alkoxyl;

R³ is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl; when X is —N(R²)—, R¹ is a substituted or unsubstituted groupselected from the group consisting of alkyl, cycloalkyl,(cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl,aralkyl, heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; orR¹ and R², together with the nitrogen to which they are attached, mayform a substituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

when X is —C(R³)(R³)—, R¹ is hydrogen, halogen, hydroxyl, sulfhydryl,nitro, cyano, or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl,acyl, acyloxy, amino, amido, azido, aminoalkyl, and alkoxyl; or the twoinstances of R³, together with the carbon to which they are attached,may form a substituted or unsubstituted 3- to 10-membered aliphatic orheterocyclic ring, wherein said ring is monocyclic, bicyclic, tricyclic,or spirocyclic; and

when X is —O—, R¹ is a substituted or unsubstituted group selected fromthe group consisting of alkyl, alkenyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl;

provided that when R⁵ is hydrogen, —XR¹ is not —N(H)CH₃, —N(H)(CH₂)₂NH₂,—N(H)(CH₂)₂COOH, —N(CH₃)₂, —N(CH₂CH₃)₂, —N(CH(CH₃)₂)₂,

An aspect of the invention is a compound represented by Formula (II) ora pharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X is —N(R²)—, —C(R³)(R³)—, or —O—;

R² is hydrogen or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, aminoalkyl, and alkoxyl;

R³ is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

when X is —N(R²)—, R¹ is a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or R¹ andR², together with the nitrogen to which they are attached, may form asubstituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

when X is —C(R³)(R³)—, R¹ is hydrogen, halogen, hydroxyl, sulfhydryl,nitro, cyano, or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl,acyl, acyloxy, amino, amido, azido, aminoalkyl, and alkoxyl; or the twoinstances of R³, together with the carbon to which they are attached,may form a substituted or unsubstituted 3- to 10-membered aliphatic orheterocyclic ring, wherein said ring is monocyclic, bicyclic, tricyclic,or spirocyclic;

when X is —O—, R¹ is a substituted or unsubstituted group selected fromthe group consisting of alkyl, alkenyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl;

R⁴ is secondary amino, tertiary amino, amido, azido, isonitrile, nitro,urea, isocyanate, carbamate, or guanidinyl; and

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl.

An aspect of the invention is a compound represented by Formula (III) ora pharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X¹ is —N(R⁶)(R⁷), —OR⁸, or —R⁹;

R⁶ and R⁷ are independently hydrogen or a substituted or unsubstitutedgroup selected from the group consisting of alkyl, cycloalkyl,(cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl,aralkyl, heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or,R⁶ and R⁷, together with the nitrogen to which they are attached, mayform a substituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

R⁸ is a substituted or unsubstituted group selected from the groupconsisting of alkyl, heteroalkyl, cycloalkyl, (cycloalkyl)alkyl,alkenyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, and aminoalkyl;

R⁹ is hydrogen, halogen, hydroxyl, sulfhydryl, or a substituted orunsubstituted group selected from the group consisting of alkyl,cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl,heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy, amino,amido, aminoalkyl, and alkoxyl;

R⁴ is secondary amino, tertiary amino, amido, azido, isonitrile, nitro,urea, isocyanate, carbamate, or guanidinyl; and

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl.

An aspect of the invention is a compound represented by Formula (IV) ora pharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X is —N(R²)—, —C(R³)(R³)—, or —O—;

R² is hydrogen or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, aminoalkyl, and alkoxyl;

R³ is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl;

when X is —N(R²)—, R¹ is a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or R¹ andR², together with the nitrogen to which they are attached, may form asubstituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

when X is —C(R³)(R³)—, R¹ is hydrogen, halogen, hydroxyl, sulfhydryl,nitro, cyano, or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl,acyl, acyloxy, amino, amido, azido, aminoalkyl, and alkoxyl; or the twoinstances of R³, together with the carbon to which they are attached,may form a substituted or unsubstituted 3- to 10-membered aliphatic orheterocyclic ring, wherein said ring is monocyclic, bicyclic, tricyclic,or spirocyclic; and

when X is —O—, R¹ is a substituted or unsubstituted group selected fromthe group consisting of alkyl, alkenyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl.

An aspect of the invention is a compound represented by Formula (V) or apharmaceutically acceptable salt thereof:

wherein R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl.

An aspect of the invention is a pharmaceutical composition, comprising acompound of the invention, or a pharmaceutically acceptable saltthereof, and a pharmaceutically acceptable carrier.

An aspect of the invention is a method of treating a fungal infection,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of the invention, thereby treating thefungal infection.

DETAILED DESCRIPTION OF THE INVENTION

Amphotericin B (AmB) is a polyene macrolide with a mycosamine appendage,the complete compound having the following structure:

AmB is generally obtained from a strain of Streptomyces nodosus. It iscurrently approved for clinical use in the United States for thetreatment of progressive, potentially life-threatening fungalinfections, including such infections as systemic or deep tissuecandidiasis, aspergillosis, cryptococcosis, blastomycosis,coccidioidomycosis, histoplasmosis, and mucormycosis, among others. Itis generally formulated for intravenous injection. Amphotericin B iscommercially available, for example, as Fungizone® (Squibb), Amphocin®(Pfizer), Abelcet® (Enzon), and Ambisome® (Astellas). Due to itsundesirable toxic side effects, dosing is generally limited to a maximumof about 1.0 mg/kg/day and total cumulative doses not to exceed about 3g in humans.

It has for many decades been widely accepted that AmB kills both yeastand human cells primarily via membrane permeabilization. However, a lackof understanding of the mechanism(s) by which AmB is toxic to yeast andhuman cells has thus far hindered the rational development of aclinically successful derivative. The longstanding accepted mechanism ofaction of AmB has been ion channel formation within a cell's membrane,leading to electrochemical gradient disruption and eventually celldeath. This model suggests that development of a less toxic derivativerequires selective ion channel formation in yeast versus human cells.

Contrary to this longstanding model, it was recently reported that theprimary mechanism of action of AmB is not ion channel formation, butsimple ergosterol binding. Gray, K C et al., Proc Natl Acd Sci USA 109:2234-9 (2012). Yeast and human cells possess different sterols,ergosterol and cholesterol, respectively. A derivative was recentlyreported in which removal of the C2′ hydroxyl group from the mycosaminesugar produced a derivative, C2′deOAmB, which surprisingly retainsergosterol-binding ability, but shows no binding to cholesterol.Wilcock, B C et al., J Am Chem Soc 135: 8488-91 (2013). Consistent withthe preferential sterol binding hypothesis, in vitro studiesdemonstrated that C2′deOAmB is toxic to yeast but not to human cells.See WO 2014/165676 to Burke et al., the entire content of which isincorporated herein by reference.

The present invention relates, at least in part, to the discovery by theinventors of further derivatives of AmB which also are characterized byimproved therapeutic index compared to AmB. The various derivatives,i.e., compounds of the invention, can be semi-synthetic or fullysynthetic.

Compounds of the invention and pharmaceutical compositions of theinvention are useful for inhibiting the growth of a fungus. In oneembodiment, an effective amount of a compound of the invention iscontacted with a fungus, thereby inhibiting growth of the fungus. In oneembodiment, a compound of the invention, or a pharmaceuticallyacceptable salt thereof, is added to or included in tissue culturemedium.

Compounds of the invention and pharmaceutical compositions of theinvention are useful for the treatment of fungal infections in asubject. In one embodiment, a therapeutically effective amount of acompound of the invention, or a pharmaceutically acceptable saltthereof, is administered to a subject in need thereof, thereby treatingthe fungal infection.

A fungus is a eukaryotic organism classified in the kingdom Fungi. Fungiinclude yeasts, molds, and larger organisms including mushrooms. Yeastsand molds are of clinical relevance as infectious agents.

Yeasts are eukaryotic organisms classified in the kingdom Fungi. Yeastsare typically described as budding forms of fungi. Of particularimportance in connection with the invention are species of yeast thatcan cause infections in mammalian hosts. Such infections most commonlyoccur in immunocompromised hosts, including hosts with compromisedbarriers to infection (e.g., burn victims) and hosts with compromisedimmune systems (e.g., hosts receiving chemotherapy or immune suppressivetherapy, and hosts infected with HIV). Pathogenic yeasts include,without limitation, various species of the genus Candida, as well as ofCryptococcus. Of particular note among pathogenic yeasts of the genusCandida are C. albicans, C. tropicalis, C. stellatoidea, C. glabrata, C.krusei, C. parapsilosis, C. guilliermondii, C. viswanathii, and C.lusitaniae. The genus Cryptococcus specifically includes Cryptococcusneoformans. Yeast can cause infections of mucosal membranes, for exampleoral, esophageal, and vaginal infections in humans, as well asinfections of bone, blood, urogenital tract, and central nervous system.This list is exemplary and is not limiting in any way.

A number of fungi (apart from yeast) can cause infections in mammalianhosts. Such infections most commonly occur in immunocompromised hosts,including hosts with compromised barriers to infection (e.g., burnvictims) and hosts with compromised immune systems (e.g., hostsreceiving chemotherapy or immune suppressive therapy, and hosts infectedwith HIV). Pathogenic fungi (apart from yeast) include, withoutlimitation, species of Aspergillus, Rhizopus, Mucor, Histoplasma,Coccidioides, Blastomyces, Trichophyton, Microsporum, andEpidermophyton. Of particular note among the foregoing are A. fumigatus,A. flavus, A. niger, H. capsulatum, C. immitis, and B. dermatitidis.Fungi can cause systemic and deep tissue infections in lung, bone,blood, urogenital tract, and central nervous system, to name a few. Somefungi are responsible for infections of the skin and nails.

Definitions

For convenience, certain terms employed in the specification, examples,and appended claims are collected here.

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

The term “acyl”, as used herein, refers to —C(═O)R, where R representsan alkyl, aryl, heteroaryl, aralkyl, or heteroaralkyl group as definedherein. Amides (RC(O)NR₂) and esters (RC(O)OR′) are classes of acylcompounds, as are ketones (RC(O)R) and aldehydes (RC(O)H). Non-limitingexamples of acyl groups include formyl, acetyl, propionyl, and benzyl.

The terms “alkenyl” and “alkynyl” are art-recognized and refer tounsaturated aliphatic groups analogous in length and possiblesubstitution to the alkyls described herein, but that contain at leastone double or triple bond, respectively.

The term “alkoxy” means an alkyl group, as defined herein, appended tothe parent molecular moiety through an oxygen atom. Representativeexamples of alkoxy include, but are not limited to, methoxy, ethoxy,propoxy, 2-propoxy, butoxy, tert-butoxy, pentyloxy, and hexyloxy.

The term “alkoxycarbonyl” means an alkoxy group, as defined herein,appended to the parent molecular moiety through a carbonyl group,represented by —C(═O)—, as defined herein. Representative examples ofalkoxycarbonyl include, but are not limited to, methoxycarbonyl,ethoxycarbonyl, and tert-butoxycarbonyl.

The term “alkyl” means a straight or branched chain hydrocarboncontaining from 1 to 10 carbon atoms. Representative examples of alkylinclude, but are not limited to, methyl, ethyl, n-propyl, iso-propyl,n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl,neopentyl, and n-hexyl.

The term “alkyl” is art-recognized, and includes saturated aliphaticgroups, including straight-chain alkyl groups, branched-chain alkylgroups, and cycloalkyl (alicyclic) groups. In certain embodiments, astraight-chain or branched-chain alkyl has about 30 or fewer carbonatoms in its backbone (e.g., C₁-C₃₀ for straight chain, C₃-C₃₀ forbranched chain), and alternatively, about 20 or fewer. In certainembodiments, a straight-chain or branched-chain alkyl has about 10 orfewer carbon atoms in its backbone. In certain embodiments, astraight-chain alkyl has 1 to 6 carbon atoms in its backbone. In certainembodiments, a branched-chain alkyl has 3 to 8 carbon atoms in itsbackbone. Representative examples of linear and branched-chain alkylgroups include, but are not limited to, methyl, ethyl, n-propyl,iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl,isopentyl, neopentyl, and n-hexyl. Cycloalkyls have from about 3 toabout 10 carbon atoms in their ring structure. In certain embodiments,cycloalkyls have 3, 4, 5, 6, or 7 carbons in the ring structure.Representative examples of cycloalkyl groups include, but are notlimited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, andcycloheptyl.

The term “alkylcarbonyl”, as used herein, means an alkyl group, asdefined herein, appended to the parent molecular moiety through acarbonyl group, as defined herein. Representative examples ofalkylcarbonyl include, but are not limited to, acetyl, 1-oxopropyl,2,2-dimethyl-1-oxopropyl, 1-oxobutyl, and 1-oxopentyl.

The term “alkylcarbonyloxy”, as used herein, means an alkylcarbonylgroup, as defined herein, appended to the parent molecular moietythrough an oxygen atom. Representative examples of alkylcarbonyloxyinclude, but are not limited to, acetyloxy, ethylcarbonyloxy, andtert-butylcarbonyloxy.

The term “alkylthio”, as used herein, means an alkyl group, as definedherein, appended to the parent molecular moiety through a sulfur atom.Representative examples of alkylthio include, but are not limited,methylthio, ethylthio, tert-butylthio, and hexylthio. The terms“arylthio”, “alkenylthio”, and “arylalkylthio,” for example, arelikewise defined in a corresponding fashion.

The term “amido”, as used herein, refers to a moiety that may berepresented by the general formula:

wherein R¹⁰ and R¹¹ each independently represent hydrogen or asubstituted or unsubstituted group selected from alkyl, cycloalkyl,cycloalkylalkyl, heterocyclyl, alkenyl, cycloalkenyl, aminoalkyl, aryl,heteroaryl, aralkyl, and heteroaralkyl. Nonlimiting examples of amidoinclude those for which R¹⁰ is hydrogen, and R¹¹ is selected frommethyl, ethyl, propyl, isopropyl, propenyl, cyclohexyl, benzyl,

Additional nonlimiting examples of amido include those for which R¹⁰ ishydrogen, and R¹¹ is selected from —CH₂NH₂, —CH₂N(CH₃)₂, and—CH(NH₂)(CH₂)_(n)NH₂, where n is an integer 1-6. Yet additionalnonlimiting examples of amido include those for which R¹⁰ is hydrogen,and R¹¹ is selected from

The terms “amino” and “amine” are art-recognized and refer to bothunsubstituted and substituted amines, e.g., a moiety that may berepresented by the general formulas:

wherein R²⁰, R²¹, and R²² each independently represent a hydrogen, analkyl, an alkenyl, —(CH₂)_(m)—R⁶¹; or R²⁰ and R²¹, taken together withthe N atom to which they are attached, complete a heterocycle havingfrom 4 to 10 atoms in the ring structure, wherein said ring ismonocyclic, bicyclic, tricyclic, or spirocyclic; R⁶¹ represents an aryl,a cycloalkyl, a cycloalkenyl, a heterocycle or a polycycle; and m iszero or an integer in the range of 1 to 8. In other embodiments, R²⁰ andR²¹ (and optionally R²²) each independently represent a hydrogen, analkyl, an alkenyl, or —(CH₂)_(m)—R⁶¹. Thus, the term “alkylamine”includes an amine group, as defined above, having a substituted orunsubstituted alkyl attached thereto, i.e., at least one of R²⁰ and R²¹is an alkyl group. Nonlimiting examples of amino groups include —NH₂,—N(H)CH₃, —N(H)CH₂CH₃, —N(H)CH₂CH₂CH₃, —N(H)CH₂CH₂CH₂CH₃, —N(CH₃)₂,—N(CH(CH₃)₂)₂, —N(CH₃)CH₂CH₃, —N(CH₃)CH₂CH₂CH₃, —N(CH₃)CH₂CH₂CH₂CH₃,—N(CH₂CH₃)₂, —N(CH₂CH₃)CH₂CH₂CH₃, —N(CH₂CH₃)CH₂CH₂CH₂CH₃,—N(CH₂CH₂CH₃)₂, —N(CH₂CH₂CH₃)CH₂CH₂CH₂CH₃, —N(CH₂CH₂CH₂CH₃)₂,

In certain embodiments, amino is —NH₂. In certain embodiments, amino is—N(H)CH₃.

The term “aminoalkyl” as used herein, means an amino group, as definedherein, appended to the parent molecular moiety through an alkyl group,also as defined herein.

The term “aromatic” refers to a planar monocyclic or polycyclicstructure characterized by a cyclically conjugated molecular moietycontaining 4n+2 electrons, wherein n is the absolute value of aninteger. Aromatic groups comprising only carbon atoms in their ringstructure are termed “aryl” groups. Aromatic groups comprising one ormore heteroatoms in their ring structure are termed “heteroaryl” or“heteroaromatic” groups. Aromatic groups containing fused, or joined,rings also are referred to as polycyclic aromatic groups. For example,bicyclic aromatic groups containing heteroatoms in a hydrocarbon ringstructure are referred to as bicyclic heteroaryl groups.

Examples of 5-, 6-, and 7-membered single-ring aromatic groups that mayinclude from zero to four heteroatoms include, for example, benzene,pyrrole, furan, thiophene, imidazole, oxazole, thiazole, triazole,pyrazole, pyridine, pyrazine, pyridazine, pyrimidine, and the like.

Non-limiting examples of polycyclic aromatic and heteroaromatic groupsinclude quinoline, isoquinoline, carbazole, naphthalene, anthracene, andpyrene.

The aryl groups of the invention can be optionally substituted with 1,2, 3, 4 or 5 substituents independently selected from the groupconsisting of alkenyl, alkoxy, alkoxycarbonyl, alkoxysulfonyl, alkyl,alkylcarbonyl, alkylcarbonyloxy, alkylsulfonyl, alkylthio, alkynyl,amido, amino, carboxy, cyano, formyl, halo, haloalkoxy, haloalkyl,hydroxyl, hydroxyalkyl, mercapto, nitro, phosphinyl, silyl and silyloxy.The term “aryl” also includes polycyclic ring systems having two or morecyclic rings in which two or more carbons are common to two adjoiningrings (the rings are “fused rings”) wherein at least one of the rings isaromatic, e.g., the other cyclic rings may be cycloalkyls,cycloalkenyls, cycloalkynyls, aryls and/or heterocyclyls.

The term “arylcarbonyloxy”, as used herein, means an arylcarbonyl group,as defined herein, appended to the parent molecular moiety through anoxygen atom. Representative examples of arylcarbonyloxy include, but arenot limited to, phenylcarbonyloxy.

The term “arylene” is art-recognized, and, as used herein, pertains to abidentate moiety obtained by removing two hydrogen atoms of an arylring, as defined above.

The term “arylalkyl” or “aralkyl”, as used herein, means an aryl group,as defined herein, appended to the parent molecular moiety through analkyl group, as defined herein.

Representative examples of arylalkyl include, but are not limited to,benzyl, 2-phenylethyl, 3-phenylpropyl, and 2-naphth-2-ylethyl.

The term “azido”, as used herein, refers to —N₃.

The term “carbamate”, as used herein, refers to a moiety that may berepresented by the general formula:

wherein R³⁰ and R³¹ each independently represent hydrogen or asubstituted or unsubstituted group selected from alkyl, cycloalkyl,cycloalkylalkyl, heterocyclyl, alkenyl, cycloalkenyl, aryl, heteroaryl,aralkyl, and heteroaralkyl. Nonlimiting examples of carbamate includethose for which R³⁰ is hydrogen, and R³¹ is selected from methyl, ethyl,propyl, isopropyl, propenyl, cyclohexyl, benzyl,

The term “carbonyl”, as used herein, means a —C(═O)— group.

The term “carboxyl”, as used herein, means a —CO₂H group.

The term “cyano”, as used herein, means a —CN group.

The term “cycloalkylalkyl” as used herein, refers to a cycloalkyl group,as defined herein, appended to the parent molecular moiety through analkyl group, also as defined herein.

The term “guanidinyl”, as used herein, refers to a moiety that may berepresented by the general formula:

wherein R⁴⁰, R⁴¹, R⁴², and R⁴³ each independently represent hydrogen ora substituted or unsubstituted group selected from alkyl, cycloalkyl,cycloalkylalkyl, heterocyclyl, alkenyl, cycloalkenyl, aryl, heteroaryl,aralkyl, and heteroaralkyl. In one embodiment, R⁴⁰, R⁴¹, R⁴², and R⁴³each represent hydrogen.

The term “halo” or “halogen” means —F, —Cl, —Br, or —I.

The term “haloalkyl” means at least one halogen, as defined herein,appended to the parent molecular moiety through an alkyl group, asdefined herein. Representative examples of haloalkyl include, but arenot limited to, chloromethyl, 2-fluoroethyl, trifluoromethyl,pentafluoroethyl, and 2-chloro-3-fluoropentyl.

The term “heteroaralkyl”, as used herein, means a heteroaryl, as definedherein, appended to the parent molecular moiety through an alkyl group,as defined herein. Representative examples of heteroarylalkyl include,but are not limited to, pyridin-3-ylmethyl and 2-(thien-2-yl)ethyl.

The term “heteroaryl”, as used herein, includes aromatic ring systems,including, but not limited to, monocyclic, bicyclic, and tricyclicrings, and have 3 to 12 atoms including at least one heteroatom, such asnitrogen, oxygen, or sulfur. For purposes of exemplification, whichshould not be construed as limiting the scope of this invention, thefollowing are examples of heteroaryl: azaindolyl, benzo(b)thienyl,benzimidazolyl, benzofuranyl, benzoxazolyl, benzothiazolyl,benzothiadiazolyl, benzotriazolyl, benzoxadiazolyl, furanyl, imidazolyl,imidazopyridinyl, indolyl, indolinyl, indazolyl, isoindolinyl,isoxazolyl, isothiazolyl, isoquinolinyl, oxadiazolyl, oxazolyl, purinyl,pyranyl, pyrazinyl, pyrazolyl, pyridinyl, pyrimidinyl, pyrrolyl,pyrrolo[2,3-d]pyrimidinyl, pyrazolo[3,4-d]pyrimidinyl, quinolinyl,quinazolinyl, triazolyl, thiazolyl, thiophenyl, tetrahydroindolyl,tetrazolyl, thiadiazolyl, thienyl, thiomorpholinyl, triazolyl ortropanyl. The heteroaryl groups may be substituted with 0, 1, 2, 3, 4 or5 substituents independently selected from alkenyl, alkoxy,alkoxycarbonyl, alkoxysulfonyl, alkyl, alkylcarbonyl, alkylcarbonyloxy,alkylsulfonyl, alkylthio, alkynyl, amido, amino, carboxy, cyano, formyl,halo, haloalkoxy, haloalkyl, hydroxyl, hydroxyalkyl, mercapto, nitro,phosphinyl, silyl and silyloxy.

The term “heteroatom” is art-recognized and refers to an atom of anyelement other than carbon or hydrogen. Illustrative heteroatoms includeboron, nitrogen, oxygen, phosphorus, sulfur, and selenium.

The term “heterocyclyl”, as used herein, refers to non-aromatic ringsystems, including, but not limited to, monocyclic, bicyclic, tricyclicand spirocyclic rings, which can be completely saturated or which cancontain one or more units of unsaturation (for the avoidance of doubt,the degree of unsaturation does not result in an aromatic ring system)and have 3 to 12 atoms including at least one heteroatom, such asnitrogen, oxygen, or sulfur. For purposes of exemplification, whichshould not be construed as limiting the scope of this invention, thefollowing are examples of heterocyclic rings: azepines, azetidinyl,morpholinyl, oxopiperidinyl, oxopyrrolidinyl, piperazinyl, piperidinyl,pyrrolidinyl, quinicludinyl, thiomorpholinyl, tetrahydropyranyl andtetrahydrofuranyl. The heterocyclyl groups may be substituted with 0, 1,2, 3, 4 or 5 substituents independently selected from alkenyl, alkoxy,alkoxycarbonyl, alkoxysulfonyl, alkyl, alkylcarbonyl, alkylcarbonyloxy,alkylsulfonyl, alkylthio, alkynyl, amido, amino, carboxy, cyano, formyl,halo, haloalkoxy, haloalkyl, hydroxyl, hydroxyalkyl, mercapto, nitro,phosphinyl, silyl and silyloxy.

The term “hydroxyl”, as used herein, means an —OH group.

The term “hydroxyalkyl”, as used herein, means at least one hydroxygroup, as defined herein, is appended to the parent molecular moietythrough an alkyl group, as defined herein.

Representative examples of hydroxyalkyl include, but are not limited to,hydroxymethyl, 2-hydroxyethyl, 3-hydroxypropyl, 2,3-dihydroxypentyl, and2-ethyl-4-hydroxyheptyl.

The term “nitro”, as used herein, means a —NO₂ group.

The term “silyl”, as used herein, includes hydrocarbyl derivatives ofthe silyl (H₃Si—) group (i.e., (hydrocarbyl)₃Si—), wherein a hydrocarbylgroups are univalent groups formed by removing a hydrogen atom from ahydrocarbon, e.g., ethyl, phenyl. The hydrocarbyl groups can becombinations of differing groups which can be varied in order to providea number of silyl groups, such as trimethylsilyl (TMS),tert-butyldiphenylsilyl (TBDPS), tert-butyldimethylsilyl (TBS/TBDMS),triisopropylsilyl (TIPS), and [2-(trimethylsilyl)ethoxy]methyl (SEM).

The term “silyloxy”, as used herein, means a silyl group, as definedherein, is appended to the parent molecule through an oxygen atom.

The term “sulfhydryl”, as used herein, means a —SH group.

The term “sulfonyl” is art-recognized and refers to —SO₂.

The term “urea”, as used herein, means a moiety that may be representedby the general formula:

wherein R⁵⁰, R⁵¹, and R⁵², each independently represent hydrogen, analkyl, an alkenyl, —(CH₂)_(m)—R⁶¹; or R⁵¹ and R⁵², taken together withthe N atom to which they are attached, complete a heterocycle havingfrom 4 to 10 atoms in the ring structure, wherein said ring ismonocyclic, bicyclic, tricyclic, or spirocyclic; R⁶¹ represents an aryl,a cycloalkyl, a cycloalkenyl, a heterocycle or a polycycle; and m iszero or an integer in the range of 1 to 8. Nonlimiting examples of ureainclude those for which R⁵⁰ is hydrogen, and R⁵¹ and R⁵² are selected inaccordance with Table 1 herein.

The definition of each expression, e.g., alkyl, m, n, and the like, whenit occurs more than once in any structure, is intended to be independentof its definition elsewhere in the same structure.

The terms triflyl, tosyl, mesyl, and nonaflyl are art-recognized andrefer to trifluoromethanesulfonyl, p-toluenesulfonyl, methanesulfonyl,and nonafluorobutanesulfonyl groups, respectively. The terms triflate,tosylate, mesylate, and nonaflate are art-recognized and refer totrifluoromethanesulfonate ester, p-toluenesulfonate ester,methanesulfonate ester, and nonafluorobutanesulfonate ester functionalgroups and molecules that contain said groups, respectively.

The abbreviations Me, Et, Ph, Tf, Nf, Ts, and Ms represent methyl,ethyl, phenyl, trifluoromethanesulfonyl, nonafluorobutanesulfonyl,p-toluenesulfonyl and methanesulfonyl, respectively. A morecomprehensive list of the abbreviations utilized by organic chemists ofordinary skill in the art appears in the first issue of each volume ofthe Journal of Organic Chemistry; this list is typically presented in atable entitled Standard List of Abbreviations.

Certain compounds contained in compositions of the invention may existin particular geometric or stereoisomeric forms. In addition, polymersof the invention may also be optically active. The inventioncontemplates all such compounds, including cis- and trans-isomers, R-and S-enantiomers, diastereomers, (D)-isomers, (L)-isomers, the racemicmixtures thereof, and other mixtures thereof, as falling within thescope of the invention. Additional asymmetric carbon atoms may bepresent in a substituent such as an alkyl group. All such isomers, aswell as mixtures thereof, are intended to be included in this invention.

If, for instance, a particular enantiomer of compound of the inventionis desired, it may be prepared by asymmetric synthesis, or by derivationwith a chiral auxiliary, where the resulting diastereomeric mixture isseparated and the auxiliary group cleaved to provide the pure desiredenantiomers. Alternatively, where the molecule contains a basicfunctional group, such as amino, or an acidic functional group, such ascarboxyl, diastereomeric salts are formed with an appropriateoptically-active acid or base, followed by resolution of thediastereomers thus formed by fractional crystallization orchromatographic means well known in the art, and subsequent recovery ofthe pure enantiomers.

It will be understood that “substitution” or “substituted with” includesthe implicit proviso that such substitution is in accordance withpermitted valence of the substituted atom and the substituent, and thatthe substitution results in a stable compound, e.g., which does notspontaneously undergo transformation such as by rearrangement,cyclization, elimination, or other reaction.

The term “substituted” is also contemplated to include all permissiblesubstituents of organic compounds. In a broad aspect, the permissiblesubstituents include acyclic and cyclic, branched and unbranched,carbocyclic and heterocyclic, aromatic and nonaromatic substituents oforganic compounds. Illustrative substituents include, for example,alkyl, alkenyl, alkynyl, aryl, aralkyl, heterocyclyl,(heterocyclyl)alkyl, (cycloalkyl)alkyl, alkoxy, aryloxy, alkoxycarbonyl,alkoxysulfonyl, aryloxycarbonyl, aryloxysulfonyl, alkylcarbonyl,arylcarbonyl, alkylcarbonyloxy, arylcarbonyloxy, alkylsulfonyl,arylsulfonyl, alkylsulfonyloxy, arylsulfonyloxy, alkylthio, arylthio,amido, amino, carboxy, cyano, formyl, halo, haloalkoxy, haloalkyl,hydroxyl, hydroxyalkyl, mercapto, nitro, phosphinyl, acyl, acyloxy,silyl and silyloxy. The permissible substituents may be one or more andthe same or different for appropriate organic compounds. For purposes ofthis invention, the heteroatoms such as nitrogen may have hydrogensubstituents and/or any permissible substituents of organic compoundsdescribed herein which satisfy the valences of the heteroatoms. Thisinvention is not intended to be limited in any manner by the permissiblesubstituents of organic compounds.

The phrase “protecting group”, as used herein, means temporarysubstituents which protect a potentially reactive functional group fromundesired chemical transformations. Examples of such protecting groupsinclude esters of carboxylic acids, silyl ethers of alcohols, andacetals and ketals of aldehydes and ketones, respectively. The field ofprotecting group chemistry has been reviewed (Greene, T.W.; Wuts, P.G.M.Protective Groups in Organic Synthesis, 2^(nd) ed.; Wiley: New York,1991). Protected forms of the inventive compounds are included withinthe scope of this invention.

For purposes of this invention, the chemical elements are identified inaccordance with the Periodic Table of the Elements, CAS version,Handbook of Chemistry and Physics, 67th Ed., 1986-87, inside cover.

Compounds of the Invention

The invention provides a number of derivatives of AmB, includingderivatives characterized by (i) certain modifications at C16; (ii) thecombination of certain modifications at C16, and certain N modificationsat C3′; (iii) the combination of other modifications at C16, and certainN modifications at C3′; (iv) the combination of certain modifications atC16, and C2′ epimerization; and (v) certain oxazolidinone derivatives.

For example, the invention provides a number of derivatives of AmB,including derivatives characterized by (i) certain urea, amide, andcarbamate modifications at C16; (ii) the combination of certain urea,amide, and carbamate modifications at C16, and certain N modificationsat C3′; (iii) the combination of certain ester, amide, aldehyde, andketone modifications at C16, and certain N modifications at C3′; (iv)the combination of certain urea, amide, and carbamate modifications atC16, and C2′ epimerization; and (v) certain oxazolidinone derivatives.

An aspect of the invention is a compound represented by Formula (I) or apharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X is —N(R²)—, —C(R³)(R³)—, or —O—;

R² is hydrogen or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, aminoalkyl, and alkoxyl;

R³ is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl;

when X is —N(R²)—, R¹ is a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or R¹ andR², together with the nitrogen to which they are attached, may form asubstituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

when X is —C(R³)(R³)—, R¹ is hydrogen, halogen, hydroxyl, sulfhydryl,nitro, cyano, or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl,acyl, acyloxy, amino, amido, azido, aminoalkyl, and alkoxyl; or the twoinstances of R³, together with the carbon to which they are attached,may form a substituted or unsubstituted 3- to 10-membered aliphatic orheterocyclic ring, wherein said ring is monocyclic, bicyclic, tricyclic,or spirocyclic; and

when X is —O—, R¹ is a substituted or unsubstituted group selected fromthe group consisting of alkyl, alkenyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl;

provided that when R⁵ is hydrogen, —XR¹ is not —N(H)CH₃, —N(H)(CH₂)₂NH₂,—N(H)(CH₂)₂COOH, —N(CH₃)₂, —N(CH₂CH₃)₂, —N(CH(CH₃)₂)₂,

In certain embodiments, R² is hydrogen.

In certain embodiments, X is —N(R²)—.

In certain embodiments, X is —N(R²)—, wherein R² is hydrogen.

In certain embodiments, X is —C(R³)(R³)—.

In certain embodiments, X is —O—.

In certain embodiments, —XR¹ is selected from the group consisting of

wherein, independently for each occurrence:

R^(a) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl;

R^(b) is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R^(c) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl; and

R^(d) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or, when—XR¹ is

R^(a) and R^(d), together with the nitrogen to which they are attached,may form a substituted or unsubstituted 3- to 10-membered heterocyclicring, wherein said ring is monocyclic, bicyclic, tricyclic, orspirocyclic.

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting ofmethyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, X is —O—; and R¹ is selected from the groupconsisting of methyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, R⁵ is hydrogen.

In certain embodiments, R⁵ is alkyl.

In certain embodiments, R⁵ is haloalkyl.

In certain embodiments of the compound of Formula (I), the—N(H)—C(O)—XR¹ moiety is replaced with —N(alkyl)-C(O)—XR¹.

An aspect of the invention is a compound represented by Formula (II) ora pharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X is —N(R²)—, —C(R³)(R³)—, or —O—;

R² is hydrogen or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, aminoalkyl, and alkoxyl;

R³ is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

when X is —N(R²)—, R¹ is a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or R¹ andR², together with the nitrogen to which they are attached, may form asubstituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

when X is —C(R³)(R³)—, R¹ is hydrogen, halogen, hydroxyl, sulfhydryl,nitro, cyano, or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl,acyl, acyloxy, amino, amido, azido, aminoalkyl, and alkoxyl; or the twoinstances of R³, together with the carbon to which they are attached,may form a substituted or unsubstituted 3- to 10-membered aliphatic orheterocyclic ring, wherein said ring is monocyclic, bicyclic, tricyclic,or spirocyclic;

when X is —O—, R¹ is a substituted or unsubstituted group selected fromthe group consisting of alkyl, alkenyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl;

R⁴ is secondary amino, tertiary amino, amido, azido, isonitrile, nitro,urea, isocyanate, carbamate, or guanidinyl; and

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl.

In certain embodiments, R² is hydrogen.

In certain embodiments, X is —N(R²)—.

In certain embodiments, X is —N(R²)—, wherein R² is hydrogen.

In certain embodiments, X is —C(R³)(R³)—.

In certain embodiments, X is —O—.

In certain embodiments, —XR¹ is selected from the group consisting of—NHCH₃,

wherein, independently for each occurrence:

R^(a) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl;

R^(b) is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R^(c) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl; and

R^(d) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or, when—XR¹ is

R^(a) and R^(d), together with the nitrogen to which they are attached,may form a substituted or unsubstituted 3- to 10-membered heterocyclicring, wherein said ring is monocyclic, bicyclic, tricyclic, orspirocyclic.

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting ofmethyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, X is —O—; and R¹ is selected from the groupconsisting of methyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, R⁴ is secondary amino.

In certain embodiments, R⁴ is tertiary amino.

In certain embodiments, R⁴ is amido.

In certain embodiments, R⁴ is azido.

In certain embodiments, R⁴ is isonitrile.

In certain embodiments, R⁴ is nitro.

In certain embodiments, R⁴ is urea.

In certain embodiments, R⁴ is isocyanate.

In certain embodiments, R⁴ is carbamate.

In certain embodiments, R⁴ is guanidinyl.

In certain embodiments, R⁴ is selected from the group consisting of

wherein

R^(a) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl.

In certain embodiments, R⁵ is hydrogen.

In certain embodiments, R⁵ is alkyl.

In certain embodiments, R⁵ is haloalkyl.

In certain embodiments of the compound of Formula (II), the—N(H)—C(O)—XR¹ moiety is replaced with —N(alkyl)-C(O)—XR¹.

An aspect of the invention is a compound represented by Formula (III) ora pharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X¹ is —N(R⁶)(R′), —OR⁸, or —R⁹;

R⁶ and R⁷ are independently hydrogen or a substituted or unsubstitutedgroup selected from the group consisting of alkyl, cycloalkyl,(cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl,aralkyl, heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or,R⁶ and R⁷, together with the nitrogen to which they are attached, mayform a substituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

R⁸ is a substituted or unsubstituted group selected from the groupconsisting of alkyl, heteroalkyl, cycloalkyl, (cycloalkyl)alkyl,alkenyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, and aminoalkyl;

R⁹ is hydrogen, halogen, hydroxyl, sulfhydryl, or a substituted orunsubstituted group selected from the group consisting of alkyl,cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl,heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy, amino,amido, aminoalkyl, and alkoxyl;

R⁴ is secondary amino, tertiary amino, amido, azido, isonitrile, nitro,urea, isocyanate, carbamate, or guanidinyl; and

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl.

In certain embodiments, R⁴ is secondary amino.

In certain embodiments, R⁴ is tertiary amino.

In certain embodiments, R⁴ is amido.

In certain embodiments, R⁴ is azido.

In certain embodiments, R⁴ is isonitrile.

In certain embodiments, R⁴ is nitro.

In certain embodiments, R⁴ is urea.

In certain embodiments, R⁴ is isocyanate.

In certain embodiments, R⁴ is carbamate.

In certain embodiments, R⁴ is guanidinyl.

In certain embodiments, R⁴ is selected from the group consisting of

wherein

R^(a) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl.

In certain embodiments, X¹ is —N(R⁶)(R′); and R⁷ is hydrogen.

In certain embodiments, X¹ is selected from the group consisting of—NHCH₃,

wherein, independently for each occurrence:

R^(a) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl;

R^(b) is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R^(c) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl; and

R^(d) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or, when X¹is

R^(a) and R^(d), together with the nitrogen to which they are attached,may form a substituted or unsubstituted 3- to 10-membered heterocyclicring, wherein said ring is monocyclic, bicyclic, tricyclic, orspirocyclic.

In certain embodiments, X¹ is selected from the group consisting of

In certain embodiments, X¹ is selected from the group consisting of

In certain embodiments, X¹ is selected from the group consisting of

In certain embodiments, X¹ is selected from the group consisting ofmethyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, X¹ is —OR⁸; and R⁸ is selected from the groupconsisting of methyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, R⁵ is hydrogen.

In certain embodiments, R⁵ is alkyl.

In certain embodiments, R⁵ is haloalkyl.

An aspect of the invention is a compound represented by Formula (IV) ora pharmaceutically acceptable salt thereof:

wherein, independently for each occurrence:

X is —N(R²)—, —C(R³)(R³)—, or —O—;

R² is hydrogen or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, acyl,amino, amido, aminoalkyl, and alkoxyl;

R³ is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl;

when X is —N(R²)—, R¹ is a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or R¹ andR², together with the nitrogen to which they are attached, may form asubstituted or unsubstituted 3- to 10-membered heterocyclic ring,wherein said ring is monocyclic, bicyclic, tricyclic, or spirocyclic;

when X is —C(R³)(R³)—, R¹ is hydrogen, halogen, hydroxyl, sulfhydryl,nitro, cyano, or a substituted or unsubstituted group selected from thegroup consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl,(heterocyclyl)alkyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl,acyl, acyloxy, amino, amido, azido, aminoalkyl, and alkoxyl; or the twoinstances of R³, together with the carbon to which they are attached,may form a substituted or unsubstituted 3- to 10-membered aliphatic orheterocyclic ring, wherein said ring is monocyclic, bicyclic, tricyclic,or spirocyclic; and

when X is —O—, R¹ is a substituted or unsubstituted group selected fromthe group consisting of alkyl, alkenyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl.

In certain embodiments, R² is hydrogen.

In certain embodiments, X is —N(R²)—.

In certain embodiments, X is —N(R²)—, wherein R² is hydrogen.

In certain embodiments, X is —C(R³)(R³)—.

In certain embodiments, X is —O—.

In certain embodiments, —XR¹ is selected from the group consisting of

wherein, independently for each occurrence:

R^(a) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl;

R^(b) is hydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy,amino, amido, azido, aminoalkyl, and alkoxyl;

R^(c) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, and aminoalkyl; and

R^(d) is hydrogen or a substituted or unsubstituted group selected fromthe group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or, when—XR¹ is

R^(a) and R^(d), together with the nitrogen to which they are attached,may form a substituted or unsubstituted 3- to 10-membered heterocyclicring, wherein said ring is monocyclic, bicyclic, tricyclic, orspirocyclic.

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting of OH

In certain embodiments, —XR¹ is selected from the group consisting of

In certain embodiments, —XR¹ is selected from the group consisting ofmethyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, X is —O—; and R¹ is selected from the groupconsisting of methyl, ethyl, propyl, isopropyl, propenyl,

In certain embodiments, R⁵ is hydrogen.

In certain embodiments, R⁵ is alkyl.

In certain embodiments, R⁵ is haloalkyl.

In certain embodiments of the compound of Formula (IV), the—N(H)—C(O)—XR¹ moiety is replaced with —N(alkyl)-C(O)—XR¹.

An aspect of the invention is a compound represented by Formula (V) or apharmaceutically acceptable salt thereof:

wherein R⁵ is selected from the group consisting of hydrogen, alkyl, andhaloalkyl.

In certain embodiments, R⁵ is hydrogen.

In certain embodiments, R⁵ is alkyl.

In certain embodiments, R⁵ is haloalkyl.

Pharmaceutical Compositions

The invention also provides pharmaceutical compositions and methods formaking same.

An aspect of the invention is a pharmaceutical composition, comprising acompound of the invention, or a pharmaceutically acceptable saltthereof, and a pharmaceutically acceptable carrier. The term“pharmaceutically acceptable carrier” means one or more compatible solidor liquid filler, diluent, or encapsulating substances which aresuitable for administration to a human or other vertebrate animal. Theterm “carrier” denotes an organic or inorganic ingredient, natural orsynthetic, with which the active ingredient is combined to facilitatethe application. The components of the pharmaceutical compositions alsoare capable of being commingled with the compounds of the presentinvention, and with each other, in a manner such that there is nointeraction which would substantially impair the desired pharmaceuticalefficacy.

In certain embodiments, the pharmaceutical composition is an intravenousdosage form.

In certain embodiments, the pharmaceutical composition is an oral dosageform.

In certain embodiments, the pharmaceutical composition is a lyophilizedpreparation of the compound with deoxycholic acid.

In certain embodiments, the pharmaceutical composition is a lyophilizedpreparation of a liposome-intercalated or liposome-encapsulated activecompound.

In certain embodiments, the pharmaceutical composition is a lipidcomplex of the compound in aqueous suspension.

In certain embodiments, the pharmaceutical composition is a cholesterylsulfate complex of the compound.

The foregoing embodiments of pharmaceutical compositions of theinvention are meant to be exemplary and are not limiting.

Also provided is a method for making such pharmaceutical compositions.The method comprises placing a compound of the invention, or apharmaceutically acceptable salt thereof, in a pharmaceuticallyacceptable carrier.

Methods of the Invention

Compounds of the invention are useful for inhibiting growth of fungi andyeast, including, in particular, fungi and yeast of clinicalsignificance as pathogens. Advantageously, the compounds of theinvention have improved therapeutic indices compared to AmB, therebyproviding agents with improved efficacy and reduced toxicity as comparedto AmB. Compounds of the invention are useful in methods of treatingfungal and yeast infections, including, in particular, systemic fungaland yeast infections. Compounds of the invention are also useful in themanufacture of medicaments for treating fungal and yeast infections,including, in particular, systemic fungal and yeast infections. Theinvention further provides the use of compounds of the invention for thetreatment of fungal and yeast infections, including, in particular,systemic fungal and yeast infections.

An aspect of the invention is a method of treating a fungal infection,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of the invention, thereby treating thefungal infection.

As used herein, “inhibit” or “inhibiting” means reduce by an objectivelymeasurable amount or degree compared to control. In one embodiment,inhibit or inhibiting means reduce by at least a statisticallysignificant amount compared to control. In one embodiment, inhibit orinhibiting means reduce by at least 5 percent compared to control. Invarious individual embodiments, inhibit or inhibiting means reduce by atleast 10, 15, 20, 25, 30, 33, 40, 50, 60, 67, 70, 75, 80, 90, or 95percent (%) compared to control.

As used herein, the terms “treat” and “treating” refer to performing anintervention that results in (a) preventing a condition or disease fromoccurring in a subject that may be at risk of developing or predisposedto having the condition or disease but has not yet been diagnosed ashaving it; (b) inhibiting a condition or disease, e.g., slowing orarresting its development; or (c) relieving or ameliorating a conditionor disease, e.g., causing regression of the condition or disease. In oneembodiment the terms “treating” and “treat” refer to performing anintervention that results in (a) inhibiting a condition or disease,e.g., slowing or arresting its development; or (b) relieving orameliorating a condition or disease, e.g., causing regression of thecondition or disease. For example, in one embodiment the terms“treating” and “treat” refer to performing an intervention that resultsin (a) inhibiting a fungal infection, e.g., slowing or arresting itsdevelopment; or (b) relieving or ameliorating a fungal infection, e.g.,causing regression of the fungal infection.

A “fungal infection” as used herein refers to an infection in or of asubject with a fungus as defined herein. In one embodiment the term“fungal infection” includes a yeast infection. A “yeast infection” asused herein refers to an infection in or of a subject with a yeast asdefined herein.

As used herein, a “subject” refers to a living mammal. In variousembodiments a subject is a non-human mammal, including, withoutlimitation, a mouse, rat, hamster, guinea pig, rabbit, sheep, goat, cat,dog, pig, horse, cow, or non-human primate. In one embodiment a subjectis a human.

As used herein, a “subject having a fungal infection” refers to asubject that exhibits at least one objective manifestation of a fungalinfection. In one embodiment a subject having a fungal infection is asubject that has been diagnosed as having a fungal infection and is inneed of treatment thereof. Methods of diagnosing a fungal infection arewell known and need not be described here in any detail.

As used herein, a “subject having a yeast infection” refers to a subjectthat exhibits at least one objective manifestation of a yeast infection.In one embodiment a subject having a yeast infection is a subject thathas been diagnosed as having a yeast infection and is in need oftreatment thereof. Methods of diagnosing a yeast infection are wellknown and need not be described here in any detail.

In certain embodiments, the compound is administered systemically.

In certain embodiments, the compound is administered parenterally.

In certain embodiments, the compound is administered intravenously.

In certain embodiments, the compound is administered intraperitoneally.

In certain embodiments, the compound is administered enterally.

In certain embodiments, the compound is administered orally.

In certain embodiments, the compound is administered intraocularly.

In certain embodiments, the compound is administered topically.

Additional routes of administration of compounds of the invention arecontemplated by the invention, including, without limitation,intravesicularly (urinary bladder), pulmonary, and intrathecally.

As used herein, the phrase “effective amount” refers to any amount thatis sufficient to achieve a desired biological effect.

As used herein, the phrase “therapeutically effective amount” refers toan amount that is sufficient to achieve a desired therapeutic effect,e.g., to treat a fungal or yeast infection.

For any compound described herein, a therapeutically effective amountcan, in general, be initially determined from in vitro studies, animalmodels, or both in vitro studies and animal models. In vitro methods arewell known and can include determination of minimum inhibitoryconcentration (MIC), minimum fungicidal concentration (MFC),concentration at which growth is inhibited by 50 percent (IC₅₀),concentration at which growth is inhibited by 90 percent (IC₉₀), and thelike. A therapeutically effective amount can also be determined fromhuman data for compounds of the invention which have been tested inhumans and for compounds which are known to exhibit similarpharmacological activities, such as other related active agents (e.g.,AmB). Higher doses may be required for parenteral administration. Theapplied dose can be adjusted based on the relative bioavailability andpotency of the administered compound. Adjusting the dose to achievemaximal efficacy based on the methods described herein and other methodsas are well-known in the art is well within the capabilities of theordinarily skilled artisan.

For any compound described herein, a therapeutically effective amountfor use in human subjects can be initially determined from in vitrostudies, animal models, or both in vitro studies and animal models. Atherapeutically effective amount for use in human subjects can also bedetermined from human data for compounds of the invention which havebeen tested in humans and for compounds which are known to exhibitsimilar pharmacological activities, such as other related active agents(e.g., AmB). Higher doses may be required for parenteral administration.The applied dose can be adjusted based on the relative bioavailabilityand potency of the administered compound. Adjusting the dose to achievemaximal efficacy based on the methods described above and other methodsas are well-known in the art is well within the capabilities of theordinarily skilled artisan.

Dosing and Formulation

Compounds of the invention can be combined with other therapeuticagents. The compound of the invention and other therapeutic agent may beadministered simultaneously or sequentially. When the other therapeuticagents are administered simultaneously, they can be administered in thesame or separate formulations, but they are administered substantiallyat the same time. The other therapeutic agents are administeredsequentially with one another and with compound of the invention, whenthe administration of the other therapeutic agents and the compound ofthe invention is temporally separated. The separation in time betweenthe administration of these compounds may be a matter of minutes or itmay be longer.

Examples of other therapeutic agents include other antifungal agents,including AmB, as well as other antibiotics, anti-viral agents,anti-inflammatory agents, immunosuppressive agents, and anti-canceragents.

As stated above, an “effective amount” refers to any amount that issufficient to achieve a desired biological effect. Combined with theteachings provided herein, by choosing among the various activecompounds and weighing factors such as potency, relativebioavailability, patient body weight, severity of adverse side-effectsand preferred mode of administration, an effective prophylactic ortherapeutic treatment regimen can be planned which does not causesubstantial unwanted toxicity and yet is effective to treat theparticular subject. The effective amount for any particular applicationcan vary depending on such factors as the disease or condition beingtreated, the particular compound of the invention being administered,the size of the subject, or the severity of the disease or condition.One of ordinary skill in the art can empirically determine the effectiveamount of a particular compound of the invention and/or othertherapeutic agent without necessitating undue experimentation. It ispreferred generally that a maximum dose be used, that is, the highestsafe dose according to some medical judgment. Multiple doses per day maybe contemplated to achieve appropriate systemic levels of compounds.Appropriate systemic levels can be determined by, for example,measurement of the patient's peak or sustained plasma level of the drug.“Dose” and “dosage” are used interchangeably herein.

Generally, daily oral doses of active compounds will be, for humansubjects, from about 0.01 milligrams/kg per day to 1000 milligrams/kgper day. It is expected that oral doses in the range of 0.5 to 50milligrams/kg, in one or several administrations per day, will yield thedesired results. Dosage may be adjusted appropriately to achieve desireddrug levels, local or systemic, depending upon the mode ofadministration. For example, it is expected that intravenousadministration would be from one order to several orders of magnitudelower dose per day. In the event that the response in a subject isinsufficient at such doses, even higher doses (or effective higher dosesby a different, more localized delivery route) may be employed to theextent that patient tolerance permits. Multiple doses per day arecontemplated to achieve appropriate systemic levels of compounds.

In one embodiment, intravenous administration of a compound of theinvention may typically be from 0.1 mg/kg/day to 20 mg/kg/day.Intravenous dosing thus may be similar to, or advantageously, may exceedmaximal tolerated doses of AmB. Intravenous dosing also may be similarto, or advantageously, may exceed maximal tolerated daily doses of AmB.Intravenous dosing also may be similar to, or advantageously, may exceedmaximal tolerated cumulative doses of AmB.

Intravenous dosing also may be similar to, or advantageously, may exceedmaximal recommended doses of AmB. Intravenous dosing also may be similarto, or advantageously, may exceed maximal recommended daily doses ofAmB. Intravenous dosing also may be similar to, or advantageously, mayexceed maximal recommended cumulative doses of AmB.

For any compound described herein the therapeutically effective amountcan be initially determined from animal models. A therapeuticallyeffective dose can also be determined from human data for compounds ofthe invention which have been tested in humans and for compounds whichare known to exhibit similar pharmacological activities, such as otherrelated active agents. Higher doses may be required for parenteraladministration. The applied dose can be adjusted based on the relativebioavailability and potency of the administered compound. Adjusting thedose to achieve maximal efficacy based on the methods described aboveand other methods as are well-known in the art is well within thecapabilities of the ordinarily skilled artisan.

The formulations of the invention are administered in pharmaceuticallyacceptable solutions, which may routinely contain pharmaceuticallyacceptable concentrations of salt, buffering agents, preservatives,compatible carriers, adjuvants, and optionally other therapeuticingredients.

Amphotericin B is commercially available in a number of formulations,including deoxycholate-based (sometimes referred to asdesoxycholate-based) formulations and lipid-based (including liposomal)formulations. Amphotericin B derivative compounds of the inventionsimilarly may be formulated, for example, and without limitation, asdeoxycholate-based formulations and lipid-based (including liposomal)formulations.

For use in therapy, an effective amount of the compound of the inventioncan be administered to a subject by any mode that delivers the compoundof the invention to the desired surface. Administering thepharmaceutical composition of the present invention may be accomplishedby any means known to the skilled artisan. Routes of administrationinclude but are not limited to oral, intravenous, intramuscular,intraperitoneal, subcutaneous, direct injection (for example, into atumor or abscess), mucosal, pulmonary (e.g., inhalation), and topical.

For intravenous and other parenteral routes of administration, thecompounds of the invention generally may be formulated similarly to AmB.For example, a compound of the invention can be formulated as alyophilized preparation with deoxycholic acid, as a lyophilizedpreparation of liposome-intercalated or -encapsulated active compound,as a lipid complex in aqueous suspension, or as a cholesteryl sulfatecomplex. Lyophilized formulations are generally reconstituted insuitable aqueous solution, e.g., in sterile water or saline, shortlyprior to administration.

For oral administration, the compounds (i.e., compounds of theinvention, and other therapeutic agents) can be formulated readily bycombining the active compound(s) with pharmaceutically acceptablecarriers well known in the art. Such carriers enable the compounds ofthe invention to be formulated as tablets, pills, dragees, capsules,liquids, gels, syrups, slurries, suspensions and the like, for oralingestion by a subject to be treated. Pharmaceutical preparations fororal use can be obtained as solid excipient, optionally grinding aresulting mixture, and processing the mixture of granules, after addingsuitable auxiliaries, if desired, to obtain tablets or dragee cores.Suitable excipients are, in particular, fillers such as sugars,including lactose, sucrose, mannitol, or sorbitol; cellulosepreparations such as, for example, maize starch, wheat starch, ricestarch, potato starch, gelatin, gum tragacanth, methyl cellulose,hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/orpolyvinylpyrrolidone (PVP). If desired, disintegrating agents may beadded, such as the cross-linked polyvinyl pyrrolidone, agar, or alginicacid or a salt thereof such as sodium alginate. Optionally the oralformulations may also be formulated in saline or buffers, e.g., EDTA forneutralizing internal acid conditions or may be administered without anycarriers.

Also specifically contemplated are oral dosage forms of the abovecomponent or components. The component or components may be chemicallymodified so that oral delivery of the derivative is efficacious.Generally, the chemical modification contemplated is the attachment ofat least one moiety to the component molecule itself, where said moietypermits (a) inhibition of acid hydrolysis; and (b) uptake into the bloodstream from the stomach or intestine. Also desired is the increase inoverall stability of the component or components and increase incirculation time in the body. Examples of such moieties include:polyethylene glycol, copolymers of ethylene glycol and propylene glycol,carboxymethyl cellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone and polyproline. Abuchowski and Davis, “SolublePolymer-Enzyme Adducts”, In: Enzymes as Drugs, Hocenberg and Roberts,eds., Wiley-Interscience, New York, N.Y., pp. 367-383 (1981); Newmark etal., J Appl Biochem 4: 185-9 (1982). Other polymers that could be usedare poly-1,3-dioxolane and poly-1,3,6-tioxocane. Preferred forpharmaceutical usage, as indicated above, are polyethylene glycolmoieties.

For the component (or derivative) the location of release may be thestomach, the small intestine (the duodenum, the jejunum, or the ileum),or the large intestine. One skilled in the art has availableformulations which will not dissolve in the stomach, yet will releasethe material in the duodenum or elsewhere in the intestine. Preferably,the release will avoid the deleterious effects of the stomachenvironment, either by protection of the compound of the invention (orderivative) or by release of the biologically active material beyond thestomach environment, such as in the intestine.

To ensure full gastric resistance a coating impermeable to at least pH5.0 is essential. Examples of the more common inert ingredients that areused as enteric coatings are cellulose acetate trimellitate (CAT),hydroxypropylmethylcellulose phthalate (HPMCP), HPMCP 50, HPMCP 55,polyvinyl acetate phthalate (PVAP), Eudragit L30D, Aquateric, celluloseacetate phthalate (CAP), Eudragit L, Eudragit S, and shellac. Thesecoatings may be used as mixed films.

A coating or mixture of coatings can also be used on tablets, which arenot intended for protection against the stomach. This can include sugarcoatings, or coatings which make the tablet easier to swallow. Capsulesmay consist of a hard shell (such as gelatin) for delivery of drytherapeutic (e.g., powder); for liquid forms, a soft gelatin shell maybe used. The shell material of cachets could be thick starch or otheredible paper. For pills, lozenges, molded tablets or tablet triturates,moist massing techniques can be used.

The therapeutic can be included in the formulation as finemulti-particulates in the form of granules or pellets of particle sizeabout 1 mm. The formulation of the material for capsule administrationcould also be as a powder, lightly compressed plugs or even as tablets.The therapeutic could be prepared by compression.

Colorants and flavoring agents may all be included. For example, thecompound of the invention (or derivative) may be formulated (such as byliposome or microsphere encapsulation) and then further contained withinan edible product, such as a refrigerated beverage containing colorantsand flavoring agents.

One may dilute or increase the volume of the therapeutic with an inertmaterial. These diluents could include carbohydrates, especiallymannitol, ca-lactose, anhydrous lactose, cellulose, sucrose, modifieddextrans and starch. Certain inorganic salts may be also be used asfillers including calcium triphosphate, magnesium carbonate and sodiumchloride. Some commercially available diluents are Fast-Flo, Emdex,STA-Rx 1500, Emcompress and Avicell.

Disintegrants may be included in the formulation of the therapeutic intoa solid dosage form. Materials used as disintegrates include but are notlimited to starch, including the commercial disintegrant based onstarch, Explotab. Sodium starch glycolate, Amberlite, sodiumcarboxymethylcellulose, ultramylopectin, sodium alginate, gelatin,orange peel, acid carboxymethyl cellulose, natural sponge and bentonitemay all be used. Another form of the disintegrants are the insolublecationic exchange resins. Powdered gums may be used as disintegrants andas binders and these can include powdered gums such as agar, Karaya ortragacanth. Alginic acid and its sodium salt are also useful asdisintegrants.

Binders may be used to hold the therapeutic agent together to form ahard tablet and include materials from natural products such as acacia,tragacanth, starch and gelatin. Others include methyl cellulose (MC),ethyl cellulose (EC) and carboxymethyl cellulose (CMC). Polyvinylpyrrolidone (PVP) and hydroxypropylmethyl cellulose (HPMC) could both beused in alcoholic solutions to granulate the therapeutic.

An anti-frictional agent may be included in the formulation of thetherapeutic to prevent sticking during the formulation process.Lubricants may be used as a layer between the therapeutic and the diewall, and these can include but are not limited to; stearic acidincluding its magnesium and calcium salts, polytetrafluoroethylene(PTFE), liquid paraffin, vegetable oils and waxes. Soluble lubricantsmay also be used such as sodium lauryl sulfate, magnesium laurylsulfate, polyethylene glycol of various molecular weights, Carbowax 4000and 6000.

Glidants that might improve the flow properties of the drug duringformulation and to aid rearrangement during compression might be added.The glidants may include starch, talc, pyrogenic silica and hydratedsilicoaluminate.

To aid dissolution of the therapeutic into the aqueous environment asurfactant might be added as a wetting agent. Surfactants may includeanionic detergents such as sodium lauryl sulfate, dioctyl sodiumsulfosuccinate and dioctyl sodium sulfonate. Cationic detergents whichcan be used and can include benzalkonium chloride and benzethoniumchloride. Potential non-ionic detergents that could be included in theformulation as surfactants include lauromacrogol 400, polyoxyl 40stearate, polyoxyethylene hydrogenated castor oil 10, 50 and 60,glycerol monostearate, polysorbate 40, 60, 65 and 80, sucrose fatty acidester, methyl cellulose and carboxymethyl cellulose. These surfactantscould be present in the formulation of the compound of the invention orderivative either alone or as a mixture in different ratios.

Pharmaceutical preparations which can be used orally include push-fitcapsules made of gelatin, as well as soft, sealed capsules made ofgelatin and a plasticizer, such as glycerol or sorbitol. The push-fitcapsules can contain the active ingredients in admixture with fillersuch as lactose, binders such as starches, and/or lubricants such astalc or magnesium stearate and, optionally, stabilizers. In softcapsules, the active compounds may be dissolved or suspended in suitableliquids, such as fatty oils, liquid paraffin, or liquid polyethyleneglycols. In addition, stabilizers may be added. Microspheres formulatedfor oral administration may also be used. Such microspheres have beenwell defined in the art. All formulations for oral administration shouldbe in dosages suitable for such administration.

For buccal administration, the compositions may take the form of tabletsor lozenges formulated in conventional manner.

For administration by inhalation, the compounds for use according to thepresent invention may be conveniently delivered in the form of anaerosol spray presentation from pressurized packs or a nebulizer, withthe use of a suitable propellant, e.g., dichlorodifluoromethane,trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide orother suitable gas. In the case of a pressurized aerosol the dosage unitmay be determined by providing a valve to deliver a metered amount.Capsules and cartridges of e.g., gelatin for use in an inhaler orinsufflator may be formulated containing a powder mix of the compoundand a suitable powder base such as lactose or starch.

Also contemplated herein is pulmonary delivery of the compounds of theinvention (or derivatives thereof). The compound of the invention (orderivative) is delivered to the lungs of a mammal while inhaling andtraverses across the lung epithelial lining to the blood stream. Otherreports of inhaled molecules include Adjei et al., Pharm Res 7:565-569(1990); Adjei et al., Int J Pharmaceutics 63:135-144 (1990) (leuprolideacetate); Braquet et al., J Cardiovasc Pharmacol 13(suppl. 5): 143-146(1989) (endothelin-1); Hubbard et al., Annal Int Med 3:206-212 (1989)(α1-antitrypsin); Smith et al., 1989, J Clin Invest 84:1145-1146(a-1-proteinase); Oswein et al., 1990, “Aerosolization of Proteins”,Proceedings of Symposium on Respiratory Drug Delivery II, Keystone,Colo., March, (recombinant human growth hormone); Debs et al., 1988, JImmunol 140:3482-3488 (interferon-gamma and tumor necrosis factor alpha)and Platz et al., U.S. Pat. No. 5,284,656 (granulocyte colonystimulating factor). A method and composition for pulmonary delivery ofdrugs for systemic effect is described in U.S. Pat. No. 5,451,569,issued Sep. 19, 1995 to Wong et al.

Contemplated for use in the practice of this invention are a wide rangeof mechanical devices designed for pulmonary delivery of therapeuticproducts, including but not limited to nebulizers, metered doseinhalers, and powder inhalers, all of which are familiar to thoseskilled in the art.

Some specific examples of commercially available devices suitable forthe practice of this invention are the Ultravent nebulizer, manufacturedby Mallinckrodt, Inc., St. Louis, Mo.; the Acorn II nebulizer,manufactured by Marquest Medical Products, Englewood, Colo.; theVentolin metered dose inhaler, manufactured by Glaxo Inc., ResearchTriangle Park, N.C.; and the Spinhaler powder inhaler, manufactured byFisons Corp., Bedford, Mass.

All such devices require the use of formulations suitable for thedispensing of compound of the invention (or derivative). Typically, eachformulation is specific to the type of device employed and may involvethe use of an appropriate propellant material, in addition to the usualdiluents, adjuvants and/or carriers useful in therapy. Also, the use ofliposomes, microcapsules or microspheres, inclusion complexes, or othertypes of carriers is contemplated. Chemically modified compound of theinvention may also be prepared in different formulations depending onthe type of chemical modification or the type of device employed.

Formulations suitable for use with a nebulizer, either jet orultrasonic, will typically comprise compound of the invention (orderivative) dissolved in water at a concentration of about 0.1 to 25 mgof biologically active compound of the invention per mL of solution. Theformulation may also include a buffer and a simple sugar (e.g., forcompound of the invention stabilization and regulation of osmoticpressure). The nebulizer formulation may also contain a surfactant, toreduce or prevent surface induced aggregation of the compound of theinvention caused by atomization of the solution in forming the aerosol.

Formulations for use with a metered-dose inhaler device will generallycomprise a finely divided powder containing the compound of theinvention (or derivative) suspended in a propellant with the aid of asurfactant. The propellant may be any conventional material employed forthis purpose, such as a chlorofluorocarbon, a hydrochlorofluorocarbon, ahydrofluorocarbon, or a hydrocarbon, including trichlorofluoromethane,dichlorodifluoromethane, dichlorotetrafluoroethanol, and1,1,1,2-tetrafluoroethane, or combinations thereof. Suitable surfactantsinclude sorbitan trioleate and soya lecithin. Oleic acid may also beuseful as a surfactant.

Formulations for dispensing from a powder inhaler device will comprise afinely divided dry powder containing compound of the invention (orderivative) and may also include a bulking agent, such as lactose,sorbitol, sucrose, or mannitol in amounts which facilitate dispersal ofthe powder from the device, e.g., 50 to 90% by weight of theformulation. The compound of the invention (or derivative) shouldadvantageously be prepared in particulate form with an average particlesize of less than 10 micrometers (μm), most preferably 0.5 to 5 μm, formost effective delivery to the deep lung.

Nasal delivery of a pharmaceutical composition of the present inventionis also contemplated. Nasal delivery allows the passage of apharmaceutical composition of the present invention to the blood streamdirectly after administering the therapeutic product to the nose,without the necessity for deposition of the product in the lung.Formulations for nasal delivery include those with dextran orcyclodextran.

For nasal administration, a useful device is a small, hard bottle towhich a metered dose sprayer is attached. In one embodiment, the metereddose is delivered by drawing the pharmaceutical composition of thepresent invention solution into a chamber of defined volume, whichchamber has an aperture dimensioned to aerosolize and aerosolformulation by forming a spray when a liquid in the chamber iscompressed. The chamber is compressed to administer the pharmaceuticalcomposition of the present invention. In a specific embodiment, thechamber is a piston arrangement. Such devices are commerciallyavailable.

Alternatively, a plastic squeeze bottle with an aperture or openingdimensioned to aerosolize an aerosol formulation by forming a spray whensqueezed is used. The opening is usually found in the top of the bottle,and the top is generally tapered to partially fit in the nasal passagesfor efficient administration of the aerosol formulation. Preferably, thenasal inhaler will provide a metered amount of the aerosol formulation,for administration of a measured dose of the drug.

The compounds, when it is desirable to deliver them systemically, may beformulated for parenteral administration by injection, e.g., by bolusinjection or continuous infusion. Formulations for injection may bepresented in unit dosage form, e.g., in ampoules or in multi-dosecontainers, with an added preservative. The compositions may take suchforms as suspensions, solutions, or emulsions in oily or aqueousvehicles, and may contain formulatory agents such as suspending,stabilizing and/or dispersing agents.

Pharmaceutical formulations for parenteral administration includeaqueous solutions of the active compounds in water-soluble form.Additionally, suspensions of the active compounds may be prepared asappropriate oily injection suspensions. Suitable lipophilic solvents orvehicles include fatty oils such as sesame oil, or synthetic fatty acidesters, such as ethyl oleate or triglycerides, or liposomes. Aqueousinjection suspensions may contain substances which increase theviscosity of the suspension, such as sodium carboxymethylcellulose,sorbitol, or dextran. Optionally, the suspension may also containsuitable stabilizers or agents which increase the solubility of thecompounds to allow for the preparation of highly concentrated solutions.

Alternatively, the active compounds may be in powder form forconstitution with a suitable vehicle, e.g., sterile pyrogen-free water,before use.

The compounds may also be formulated in rectal or vaginal compositionssuch as suppositories or retention enemas, e.g., containing conventionalsuppository bases such as cocoa butter or other glycerides.

In addition to the formulations described above, the compounds may alsobe formulated as a depot preparation. Such long acting formulations maybe formulated with suitable polymeric or hydrophobic materials (forexample as an emulsion in an acceptable oil) or ion exchange resins, oras sparingly soluble derivatives, for example, as a sparingly solublesalt.

The pharmaceutical compositions also may comprise suitable solid or gelphase carriers or excipients. Examples of such carriers or excipientsinclude but are not limited to calcium carbonate, calcium phosphate,various sugars, starches, cellulose derivatives, gelatin, and polymerssuch as polyethylene glycols.

Suitable liquid or solid pharmaceutical preparation forms are, forexample, aqueous or saline solutions for inhalation, microencapsulated,encochleated, coated onto microscopic gold particles, contained inliposomes, nebulized, aerosols, pellets for implantation into the skin,or dried onto a sharp object to be scratched into the skin. Thepharmaceutical compositions also include granules, powders, tablets,coated tablets, (micro)capsules, suppositories, syrups, emulsions,suspensions, creams, drops or preparations with protracted release ofactive compounds, in whose preparation excipients and additives and/orauxiliaries such as disintegrants, binders, coating agents, swellingagents, lubricants, flavorings, sweeteners or solubilizers arecustomarily used as described above. The pharmaceutical compositions aresuitable for use in a variety of drug delivery systems. For a briefreview of methods for drug delivery, see Langer R, Science 249:1527-33(1990), which is incorporated herein by reference.

The compounds of the invention and optionally other therapeutics may beadministered per se (neat) or in the form of a pharmaceuticallyacceptable salt. When used in medicine the salts should bepharmaceutically acceptable, but non-pharmaceutically acceptable saltsmay conveniently be used to prepare pharmaceutically acceptable saltsthereof. Such salts include, but are not limited to, those prepared fromthe following acids: hydrochloric, hydrobromic, sulphuric, nitric,phosphoric, maleic, acetic, salicylic, p-toluene sulphonic, tartaric,citric, methane sulphonic, formic, malonic, succinic,naphthalene-2-sulphonic, and benzene sulphonic. Also, such salts can beprepared as alkaline metal or alkaline earth salts, such as sodium,potassium or calcium salts of the carboxylic acid group.

Suitable buffering agents include: acetic acid and a salt (1-2% w/v);citric acid and a salt (1-3% w/v); boric acid and a salt (0.5-2.5% w/v);and phosphoric acid and a salt (0.8-2% w/v). Suitable preservativesinclude benzalkonium chloride (0.003-0.03% w/v); chlorobutanol (0.3-0.9%w/v); parabens (0.01-0.25% w/v) and thimerosal (0.004-0.02% w/v).

Pharmaceutical compositions of the invention contain an effective amountof a compound of the invention and optionally at least one additionaltherapeutic agent included in a pharmaceutically acceptable carrier.

The therapeutic agent(s), including specifically but not limited to thecompound of the invention, may be provided in particles. Particles asused herein means nanoparticles or microparticles (or in some instanceslarger particles) which can consist in whole or in part of the compoundof the invention or the other therapeutic agent(s) as described herein.The particles may contain the therapeutic agent(s) in a core surroundedby a coating, including, but not limited to, an enteric coating. Thetherapeutic agent(s) also may be dispersed throughout the particles. Thetherapeutic agent(s) also may be adsorbed into the particles. Theparticles may be of any order release kinetics, including zero-orderrelease, first-order release, second-order release, delayed release,sustained release, immediate release, and any combination thereof, etc.The particle may include, in addition to the therapeutic agent(s), anyof those materials routinely used in the art of pharmacy and medicine,including, but not limited to, erodible, nonerodible, biodegradable, ornonbiodegradable material or combinations thereof. The particles may bemicrocapsules which contain the compound of the invention in a solutionor in a semi-solid state. The particles may be of virtually any shape.

Both non-biodegradable and biodegradable polymeric materials can be usedin the manufacture of particles for delivering the therapeutic agent(s).Such polymers may be natural or synthetic polymers. The polymer isselected based on the period of time over which release is desired.Bioadhesive polymers of particular interest include bioerodiblehydrogels described in Sawhney H S et al. (1993) Macromolecules26:581-7, the teachings of which are incorporated herein. These includepolyhyaluronic acids, casein, gelatin, glutin, polyanhydrides,polyacrylic acid, alginate, chitosan, poly(methyl methacrylates),poly(ethyl methacrylates), poly(butylmethacrylate), poly(isobutylmethacrylate), poly(hexylmethacrylate), poly(isodecyl methacrylate),poly(lauryl methacrylate), poly(phenyl methacrylate), poly(methylacrylate), poly(isopropyl acrylate), poly(isobutyl acrylate), andpoly(octadecyl acrylate).

The therapeutic agent(s) may be contained in controlled release systems.The term “controlled release” is intended to refer to anydrug-containing formulation in which the manner and profile of drugrelease from the formulation are controlled. This refers to immediate aswell as non-immediate release formulations, with non-immediate releaseformulations including but not limited to sustained release and delayedrelease formulations. The term “sustained release” (also referred to as“extended release”) is used in its conventional sense to refer to a drugformulation that provides for gradual release of a drug over an extendedperiod of time, and that preferably, although not necessarily, resultsin substantially constant blood levels of a drug over an extended timeperiod. The term “delayed release” is used in its conventional sense torefer to a drug formulation in which there is a time delay betweenadministration of the formulation and the release of the drug therefrom. “Delayed release” may or may not involve gradual release of drugover an extended period of time, and thus may or may not be “sustainedrelease.”

Use of a long-term sustained release implant may be particularlysuitable for treatment of chronic conditions. “Long-term” release, asused herein, means that the implant is constructed and arranged todeliver therapeutic levels of the active ingredient for at least 7 days,and preferably 30-60 days. Long-term sustained release implants arewell-known to those of ordinary skill in the art and include some of therelease systems described above.

Having now described the present invention in detail, the same will bemore clearly understood by reference to the following examples, whichare included herewith for purposes of illustration only and are notintended to be limiting of the invention.

EXAMPLES Example 1. C₁₅-C₁₆ Oxazolidinone 1

Preparation of 1-1: Step 1: A 2 L round-bottom flask was charged withAmphotericin B (Chem-Impex International) (I) (50 g, ca. 54 mmol). Amixture of DMF/MeOH (900 mL/450 mL) was added, followed by pyridine (25mL) and FMOC-OSu (27.4 g, 81.3 mmol). The mixture was stirred at RT for12 h and then poured into ether (5 L). The yellow precipitate wascollected by filtration in a fritted glass funnel. It was then washedwith more ether (4 L) and dried under high vacuum (covered with aluminumfoil to prevent exposure to light) overnight. The yellow solid (64g, >100% yield) thus obtained was used in the next step without furtherpurification. LCMS: Observed mass 744 m/z, The [M+H]⁺ ion was notobserved.

Step 2: A 2 L round-bottom flask was charged with FMOC-protectedAmphotericin B (64 g). A mixture of THF/MeOH (700 mL/700 mL) was added.The mixture was cooled in an ice/water bath and stirred under N₂ for 30min. Camphor-10-sulfonic acid (CSA) (3.8 g) was added in one portion.The mixture was stirred at 0° C. for 2 h. Triethylamine (8 mL) was thenadded. The mixture was concentrated to approximately half its originalvolume and then poured into a mixture of hexanes/ether (2 L/2 L). Themixture was stirred at RT for 15 min and the yellow precipitate wascollected by filtration using a fritted glass funnel. The solid waswashed with more ether (ca. 500 mL) and dried under high vacuum (coveredwith aluminum foil to prevent exposure to light) for 2 h. The yellowsolid thus obtained (68 g, >100% yield) was used in the next stepwithout further purification. LCMS, Observed mass 742.7 m/z, the [M+H]⁺ion was not observed.

Preparation of 1-2: A 1 L round-bottom flask was charged withFMOC-protected ketal of amphotericin B 1-1 (68 g, ca. 55 mmol).Anhydrous THF (500 mL) was added and the suspension was stirred at RTfor 10 min. Triethylamine (20 mL, 143.5 mmol) was then added. Themixture was stirred at RT for a further 15 min. Diphenylphosphoryl azide(16 mL, 68.6 mmol) was added in four equal portions at 3 minuteintervals via syringe. The mixture was then heated to 50° C. and stirredfor 2 hours. The reaction was cooled to room temperature and then pouredinto MTBE (1 L). The yellow precipitate was collected by filtrationusing a fritted glass funnel and then mixed with silica gel (ca. 100 g)and treated with DCM/MeOH (50 mL/5 mL). The slurry was concentrated,loaded onto a silica gel column (10 cm×48 cm) and purified using alinear gradient of 0-10% MeOH/DCM collecting 50 mL fractions. Purefractions (R_(f)=0.5 on TLC, 10% MeOH/CH₂C12) were combined andconcentrated in vacuo to afford 1-2 as a yellow/orange solid (19.5 g,16.86 mmol, 30.6% yield).

Preparation of 1-3: Compound 1-2 (300 milligrams) was dissolved in THFand treated with 16% formic acid in water. The solution was heated at50° C. for 2 h. Evaporation of the solvent and purification byreverse-phase HPLC provided 1-3 (23 milligrams) (LCMS, 1166.1, M+Na).

Preparation of 1: Treatment of 1-3 with 2 equivalents of DMAP in DMSO atRT for 2 h or trimethylamine in DMF at RT for 12 h, followed bypurification by RP-HPLC and lyophilization, provides the target compound1.

Example 2. C16 Ureas 2

Compounds 2 can also be prepared according to the method described inScheme 2, route 2. Isocyanate 3-2, prepared as described below, istreated with an amine (5-50 equivalents) in THF (0.1-0.6 M) attemperatures ranging from 23° C. to 80° C. Removal of the silyl groupswith HF/pyridine and deketalization/purification with 0.3% formic acidin DMSO followed by purification in H₂O/CH₃CN mixtures using 0.3% formicacid as a modifier provides the target ureas.

Specific Compounds 2

where H—NR₁R₂ is defined in Table 1:

TABLE 1

A

B

C

D

E

F

G

H

I

J

K

L

M

N

O

P

Q

R

S

T

U

V

W

X

Y

Z

AA

AB

AC

AD

AE

AF

AG

AH

AI

AJ

AK

AL

AM

AN

AO

AP

AQ

AR

AS

AT

AU

AV

AW

AX

AY

AZ

BA

BB

BC

BD

BE

BF

BG

BH

BI

BJ

BK

BL

BM

BN

BO

BP

BQ

BR

BS

BT

BU

BV

BW

BX

BY

BZ

CA

CB

CC

CD

CE

CF

CG

CH

CI

CJ

CK

CL

CM

CN

CO

CP

CQ

CR

CS

CT

CU

Example 3. Preparation of Compound 2-I

Preparation of 2-2: To a solution of 1-2 (Example 1; 350 mg, 302.4 μmol,1.00 eq.) in THF (16 mL) was added piperazine (93 mg, 1.08 mmol, 3.57eq.) at 15° C. The mixture was stirred at 50° C. for 2 h under Ar₂. Ayellow precipitate formed, and HPLC showed that 1-2 was consumed. Themixture was poured into MTBE (350 mL), and the solid was collected byfiltration to provide crude 2-2-I (450 mg). The filter cake washed withEtOAc/MeOH=1:1 (3 mL), filtered, give 1-2 as yellow solid.

Preparation of 2: A solution of 2-2-I (450 mg) in aqueous formic acid(16% v/v, 3 mL) was stirred at 45° C. for 20 min. HPLC showed the methylketal was hydrolyzed completely. Toluene (20 mL) was added, and themixture was concentrated in vacuum at 40° C. The residue was dissolvedin DMSO (5 mL) and purified by prep-HPLC (C18, 5-μm, 250×50 mm, 80mL/min, 3% to 33% MeCN: 0.1% FA (aq) over 20 minutes) to provide 2-I(70.00 mg, 23% yield) as a yellow solid.

¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ 8.70 (s, 1H) 6.16-6.52 (m, 14H)5.37-5.55 (m, 2H) 4.80 (s, 1H) 4.50 (br. s., 1H) 4.39 (m, 1H) 4.25-4.37(m, 4H) 3.37-3.61 (m, 10H) 3.25-3.37 (m, 1H) 2.97 (s, 4H) 2.37-2.42 (m,3H) 2.24 (s, 2H) 1.47-1.87 (m, 14H) 1.29 (d, J=6 Hz, 3H) 1.23 (d, J=6Hz, 3H) 1.12 (d, J=6 Hz, 3H) 1.11 (d, J=6 Hz, 3H). LCMS (ESI): m/z:[M+Na] calcd for C₅₁H₈₂N₄O₁₆Na: 1029.57; found 1029.6.

Example 4. Preparation of Compound 2-BF

Step 1: A 1 L round-bottom flask was charged with 1-2 (Example 1; 11.36g, ca. 9.8 mmol). Anhydrous THF (100 mL) was added and thesuspension/solution was stirred at RT for 10 min. Methyl amine (40 mL, 2M in THF, 80 mmol) was then added and the mixture was stirred at RT for12 h. More methyl amine (10 mL, 2 M in THF, 20 mmol) was added and themixture was further stirred for 15 h until most of the starting materialwas consumed as determined by LCMS analysis (Method 1). A small amount(1-2%) of the starting Fmoc-deprotected oxazolidinone (RT=4.1 min)remained when the reaction was stopped. The reaction was then dilutedwith MTBE (500 mL) and filtered through a fritted glass funnel. Thesolid was washed with MTBE (50 mL) and then dried under high vacuum for1 h. A yellow/orange solid, 2-2-BF, (9.9 g, >100% yield) was obtained.LCMS, RT=3.78 min, 966.8 m/z [M+H]⁺.

Step 2: 2-2-BF was dissolved in 6 mL DMSO and diluted with water (1 mL).The pH of the solution was adjusted to 3 with 20% aq. formic acid. Themixture was loaded on an HPLC column and subjected to HPLC purification.Column: Microsorb (100 Å pore size, 10 μm particle size) C-18 column(50×450 mm); flow rate=100 mL/min; mobile phase A: 99.7% water, 0.3%HCOOH; mobile phase B: 99.7% ACN, 0.3% HCOOH; gradient elution from 0% Bto 95% B over 95 min; detection at 383 nm. The total volume of eluantwas 10 L. The compound eluted at 31-35% of buffer B. Eight 50 mLfractions containing the desired compound were combined and evaporatedunder reduced pressure at a bath temperature between 30-40° C. to 20% ofthe initial volume. The pH of the solution was adjusted to 7.5 withsodium bicarbonate. The suspension (100 mL) thus obtained wascentrifuged at 4000 rpm. The supernatant was separated and the solidportion re-suspended in water (100 mL) and centrifuged again. Theprocedure was repeated three times until the disappearance of the saltsignal on the ELSD chromatogram. The final solid was re-suspended inwater and subjected to lyophilization to afford the target material(2-BF, 760 mg, 38%) as a yellow powder.

Example 5. Preparation of Compound 2-BC

Step 1: To a 20 mL vial was added b-alanine allylester hydrochloride(1.125 g, 6.79 mmol, 39 eq.), sodium carbonate (2.19 g, 20.66 mmol, 120eq.), and DMF (8.6 mL). The resulting suspension was stirred at roomtemperature for 15 minutes. The suspension was then filtered throughCelite followed by filtration through a syringe tip 0.2-μm filter. Theresulting b-alanine allylester free base was then added to a 20 mL vialcontaining 1-2 (200 mg, 0.174 mmol, 1 eq.). The reaction was placed in apreheated heating block at 40° C. and allowed to stir for 5 h. Thereaction was then directly purified directly by prep HPLC (C18, 5-mm,30×150 mm, 25 mL/min, 95:5 to 40:60 0.3% HCO2H (aq):MeCN over 10minutes). Upon removal of the acetonitrile and aqueous formic acidsolution in vacuo at 35° C., the C-13 methyl ketal is converted to ahemiketal yielding 2-BC-allylester as a yellow solid (59.4 mg, 32.5%yield).

Step 2: To a 40 mL vial was added 2-BC-allylester (370 mg, 352.3 mmol, 1eq.), and thiosalicylic acid (203.4 mg, 1.76 mmol, 5 eq.). The vial wasthen brought into a glovebox and Pd(PPh₃)₄ was added (205 mg, 0.18 mmol,0.5 eq.). The vial was sealed with a septa cap, removed from theglovebox, and DMF was added (17.6 mL, 0.2 M) via syringe. The reactionthen stirred at room temperature for 1 h. The reaction was then pouredinto Et₂O (370 mL) in multiple 50 mL centrifuge tubes. The resultingsuspension was then centrifuged at 3700 G for 5 minutes. The pale redsupernatant was decanted and the resulting yellow/orange solid wasdissolved in DMSO and purified by prep HPLC (C18, 5-mm, 50×250 mm, 80mL/min, 80:20 to 40:60 0.3% HCO2H (aq):MeCN over 9 minutes) yielding2-BC as a yellow solid (124.4 mg, 35% yield). ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 6.45 (br. s., 11H), 5.02 (s, 1H), 4.42-4.84(m, 5H), 4.31 (br. s., 1H), 3.68-4.09 (m, 7H), 3.40 (d, J=9.26 Hz, 1H),2.47-2.82 (m, 5H), 2.40 (d, J=14.11 Hz, 2H), 2.24 (d, J=6.17 Hz, 2H),1.60-2.16 (m, 11H), 1.44-1.60 (m, 5H), 1.40 (d, J=6.62 Hz, 3H), 1.27 (d,J=6.17 Hz, 3H), 1.21 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcdfor C₅₀H₇₉N₈O₁₇Na: 1033.5; found 1033.4.

Example 6. Synthesis of Compound 2-B

Compound 2-B was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with N,N-Dimethylamine.¹H NMR (400 MHz, Methanol-d4+Py-d⁵): δ 8.82 (s, 1H) 6.17-6.54 (m, 14H)5.52 (d, 1H) 4.80 (s, 1H) 4.52 (br. s., 1H) 4.40 (s, 1H) 4.30-4.38 (m,2H) 4.28-4.30 (m, 2H) 3.78-3.87 (m, 3H) 3.78 (t, J=4.8 Hz, 3H) 3.40-3.42(m, 1H) 3.26-3.28 (m, 2H) 2.80 (s, 6H) 2.43-2.247 (m, 3H) 2.27-2.23 (m,2H) 1.57-1.90 (m, 13H) 1.33 (d, J=6 Hz, 3H) 1.25 (d, J=6 Hz, 3H) 1.14(d, J=6 Hz, 3H) 1.03 (d, J=6 Hz, 3H). LCMS (ESI): m/z: [M+Na] calked forC₄₉H₇₉N₃O₁₆Na: 988.55; found 988.6.

Example 7. Synthesis of Compound 2-C

Compound 2-C was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with Piperidine. ¹H NMR(400 MHz, Methanol-d4+Py-d⁵): δ 8.67 (s, 1H) 6.16-6.50 (m, 14H)5.36-5.40 (m, 2H) 4.79 (s, 1H) 4.63 (br. s., 1H) 4.51 (s, 1H) 4.27-4.37(m, 4H) 3.66-3.84 (m, 7H) 3.25 (m, 3H) 2.23-2.42 (m, 4H) 1.53-1.90 (m,22H) 1.33 (d, J=6 Hz, 3H) 1.23 (d, J=6 Hz, 3H) 1.12 (d, J=6 Hz, 3H) 1.03(d, J=6 Hz, 3H). LCMS (ESI): m/z: [M+Na] calked for C52H₈₃N₃O₁₆Na:1028.58; found 1028.6.

Example 8. Synthesis of Compound 2-J

Compound 2-J was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with N-methylpiperazine.¹H NMR (400 MHz, Methanol-d4+Py-d⁵): δ 8.71 (s, 1H) 6.18-6.54 (m, 14H)5.42-5.52 (m, 2H) 4.84 (s, 1H) 4.66 (br. s., 1H) 4.41 (s, 1H) 4.29-4.38(m, 5H) 3.37-3.85 (m, 4H) 3.26-3.47 (m, 7H) 2.29-2.43 (m, 3H) 2.25-2.26(m, 6H) 2.09 (s, 4H) 1.58-1.60 (m, 15H) 1.32 (d, J=6 Hz, 3H) 1.26 (d,J=6 Hz, 3H) 1.24 (d, J=6 Hz, 3H) 1.13 (d, J=6 Hz, 3H). LCMS (ESI): m/z:[M+Na] calcd for C₅₂H₈₄N₄O₁₆Na: 1043.59; found 1043.5.

Example 9. Synthesis of Compound 2-D

Compound 2-D was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with azetidine. ¹H NMR(400 MHz, Methanol-d4+Pyr-d5): δ 8.77 (s, 1H) 6.16-6.55 (m, 14H)4.84-5.52 (m, 2H) 4.84 (s, 1H) 4.66 (br. s., 1H) 4.53 (t, 1H) 4.24-4.36(m, 5H) 3.86-3.88 (m, 5H) 3.70-3.85 (m, 3H) 3.28-3.50 (m, 4H) 2.43-2.45(m, 3H) 2.26 (s, 2H) 1.59-1.92 (m, 14H) 1.35 (d, J=6 Hz, 3H) 1.26 (d,J=6 Hz, 3H) 1.15 (d, J=6 Hz, 3H) 1.07 (d, J=6 Hz, 3H). LCMS (ESI): m/z:[M+Na] calcd for C₅₀H₇₉N₃O₁₆Na: 1000.55; found 1000.5.

Example 10. Synthesis of Compound 2-F

Compound 2-F was synthesized in the manner similar to Compound 2-I,except piperazine was substituted withN′,N′1-dimethylethane-1,2-diamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5):δ 8.62 (s, 1H) 6.17-6.51 (m, 14H) 5.34-5.48 (m, 2H) 4.77 (s, 1H) 4.48(br. s., 1H) 4.36 (m, 1H) 4.23-4.30 (m, 3H) 4.05 (m, 1H) 3.42-3.81 (m,6H) 3.22-3.26 (m, 1H) 2.92 (m, 2H) 2.620 (s, 6H) 2.44-2.52 (m, 4H)2.20-2.44 (m, 2H) 1.58-1.86 (m, 6H) 1.51-1.54 (m, 7H) 1.32 (d, J=6 Hz,3H) 1.23 (d, J=6 Hz, 3H) 1.12 (d, J=6 Hz, 3H) 1.04 (d, J=6 Hz, 3H). LCMS(ESI): m/z: [M+H] calcd for C₅₁H₈₄N₄O₁₆: 1009.59; found 1009.6.

Example 11. Synthesis of Compound 2-T

Compound 2-T was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-(pyridin-2-yl)ethanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ 8.64(s, 1H) 8.38-8.39 (m, 1H) 7.53-7.57 (m, 1H) 7.28-7.31 (m, 1H) 7.17-7.20(m, 14H) 6.16-6.35 (m, 14H) 5.33-5.50 (m, 3H) 4.84 (s, 1H) 4.64 (br. s.,2H) 4.49 (t, 1H) 3.71-4.44 (m, 3H) 3.57-3.68 (m, 2H) 3.54-3.55 (m, 3H)3.45-3.54 (m, 2H) 3.23-3.25 (m, 1H) 2.96-2.97 (m, 2H) 1.85-2.23 (m, 4H)1.82-1.85 (m, 2H) 1.50-1.82 (m, 14H) 1.32 (d, J=6 Hz, 3H) 1.23 (d, J=6Hz, 3H) 1.12 (d, J=6 Hz, 3H) 1.04 (d, J=6 Hz, 3H). LCMS (ESI): m/z:[M+Na] calcd for C₅₄H₈₂N₄O₁₆Na: 1065.57; found 1065.4.

Example 12. Synthesis of Compound 2-U

Compound 2-U was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withpyridin-2-ylmethanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ 8.80(s, 1H) 8.42-8.43 (m, 1H) 7.51-7.52 (m, 1H) 7.38-7.40 (m, 1H) 7.02-7.03(m, 1H) 6.21-6.58 (m, 14H) 5.56 (s, 1H) 4.91 (s, 1H) 4.23-4.71 (m, 5H)3.77-3.89 (m, 4H) 3.25-3.50 (m, 4H) 2.30-3.25 (m, 1H) 2.28-2.30 (m, 2H)1.92-2.28 (m, 2H) 1.57-1.89 (m, 13H) 1.35 (d, J=6 Hz, 3H) 1.26 (d, J=6Hz, 3H) 1.14 (d, J=6 Hz, 3H) 1.09 (d, J=6 Hz, 3H). LCMS (ESI): m/z:[M+Na] calcd for C₅₃H₅₀N₄O₁₆Na: 1051.56; found 1051.4.

Example 13. Synthesis of Compound 2-Y

Compound 2-Y was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(S)-1-aminopropan-2-ol. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ 8.74 (s,1H), 6.15-6.51 (m, 14H), 5.29-5.37 (m, 2H), 4.85 (s, 1H), 4.65 (br. s.,1H), 4.38 (t, 1H), 4.33-4.35 (m, 5H), 3.85-3.87 (m, 2H), 3.71-3.74 (m,1H), 3.66-3.69 (m, 1H), 3.23-3.49 (m, 2H), 2.21-2.37 (m, 3H), 1.55-1.87(m, 11H), 1.23-1.31 (m, 13H), 1.06-1.13 (m, 15H). LCMS (ESI): m/z:[M+Na] calcd for C₅₀H₅₁N₃O₁₇Na: 1018.56; found 1018.5.

Example 14. Synthesis of Compound 2-AB

Compound 2-AB was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with1-(piperazin-1-yl)ethanone. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ8.68-8.71 (s, 2H) 6.21-6.53 (m, 13H) 5.54 (m, 2H) 4.69-4.84 (m, 5H)4.35-4.40 (m, 2H) 3.67-3.91 (m, 9H) 3.38-3.53 (m, 4H) 1.59-2.50 (m, 35H)1.30 (d, J 30=6 Hz, 3H) 1.26 (d, J=6 Hz, 3H) 1.20 (d, J=6 Hz, 3H) 1.10(d, J=6 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd for C₅₃H₈₄N₄O₁₆Na:1071.58.

Example 15. Synthesis of Compound 2-AF

Compound 2-AF was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-morpholinoethanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ 8.62 (s,1H) 6.18-6.52 (m, 14H) 5.39-5.51 (m, 2H) 4.86 (s, 1H) 4.68 (br. s., 1H)4.46-4.52 (t, 1H) 4.35 (m, 2H) 3.73-3.88 (m, 4H) 3.37-3.61 (m, 10H)2.26-2.48 (m, 12H) 1.34-1.88 (m, 12H) 1.25 (d, J=6 Hz, 3H) 1.14 (d, J=6Hz, 3H) 1.07 (d, J=6 Hz, 3H) 1.06 (d, J=6 Hz, 3H). LCMS (ESI): m/z:[M+H] calcd for C₅₃H₈₇N₄O₁₇: 1051.60; found 1051.70.

Example 16. Synthesis of Compound 2-BN

Compound 2-BN was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-(4-methylpiperazin-1-yl)ethanamine. ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 6.03-6.60 (m, 14H) 5.49 (d, J=6.02 Hz, 1H)5.35-5.43 (m, 1H) 4.80 (s, 1H) 4.65 (d, J=6.90 Hz, 1H) 4.46-4.54 (m, 1H)4.33 (br. s., 1H) 4.19-4.28 (m, 2H) 4.01-4.12 (m, 2H) 3.79-3.88 (m, 2H)3.72 (d, J=11.17 Hz, 1H) 3.51-3.68 (m, 3H) 3.43 (dd, J=9.03, 6.02 Hz,2H) 3.27 (d, J=9.79 Hz, 2H) 2.32-2.59 (m, 13H) 2.16-2.28 (m, 5H)1.81-2.04 (m, 5H) 1.36-1.80 (m, 11H) 1.34 (d, J=6.15 Hz, 3H) 1.25 (d,J=6.40 Hz, 3H) 1.15 (d, J=6.27 Hz, 3H) 1.06 (d, J=7.03 Hz, 3H). LCMS(ESI): m/z: [M+H] calcd for C₅₄H₉₀N₅O₁₆: 1064.63; found 1064.6.

Example 17. Synthesis of Compound 2-BO

Compound 2-BO was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withcyclopropylmethanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ5.86-6.26 (m, 13H), 5.22 (d, J=5.73 Hz, 1H), 5.08 (br. s., 2H), 4.33(br. s., 1H), 4.21 (br. s., 1H), 3.92-4.11 (m, 2H), 3.68-3.86 (m, 2H),3.53 (t, J=9.04 Hz, 1 H), 3.28-3.46 (m, 2H), 3.01-3.15 (m, 3H), 2.93 (d,J=8.82 Hz, 1H), 2.65-2.81 (m, 2H), 2.25-2.50 (m, 2H), 2.01-2.19 (m, 2H),1.85-1.97 (m, 2H), 1.03-2.00 (m, 14H), 1.01 (d, J=5.73 Hz, 3H), 0.92 (d,J=6.17 Hz, 3H), 0.81 (d, J=6.17 Hz, 3H), 0.73 (d, J=7.06 Hz, 3H). LCMS(ESI): m/z: [M+Na] calcd for C₅₁H₈₁N₃O₁₆Na: 1014.56; found 1014.6.

Example 18. Synthesis of Compound 2-BP

Compound 2-BP was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(2-chlorophenyl)methanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm8.73-8.85 (m, 1H) 7.47-7.54 (m, 1H) 7.21-7.25 (m, 1H) 7.10-7.15 (m, 1H)7.01-7.09 (m, 1H) 6.10-6.58 (m, 12H) 5.49-5.59 (m, 1H) 5.36-5.41 (m, 1H)4.81 (s, 1H) 4.62-4.71 (m, 1H) 4.48 (s, 4H) 4.31-4.41 (m, 2H) 4.22-4.28(m, 1H) 4.06-4.17 (m, 1H) 3.80-3.91 (m, 1H) 3.67-3.78 (m, 2H) 3.54-3.64(m, 1H) 3.37-3.48 (m, 1H) 3.18-3.27 (m, 1H) 2.98-3.09 (m, 1H) 2.16-2.64(m, 5H) 1.99-2.13 (m, 1H) 1.79-1.96 (m, 3H) 1.40-1.78 (m, 7H) 1.32 (d,J=6.17 Hz, 4H) 1.24 (d, J=6.17 Hz, 3H) 1.12 (d, J=6.62 Hz, 3H) 1.05 (d,J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd for C₅₄H₈₀ClN₃O₁₆Na:1084.52; found 1084.4.

Example 19. Synthesis of Compound 2-BQ

Compound 2-BQ was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(3-chlorophenyl)methanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm8.76-8.82 (m, 1H) 7.33 (s, 2H), 7.08-7.16 (m, 3H), 6.86-6.93 (m, 1H),6.07-6.63 (m, 12H), 5.48-5.58 (m, 1H), 5.36-5.41 (m, 1H), 4.81 (s, 1H),4.62-4.74 (m, 1H), 4.53 (t, J=10.36 Hz, 1H), 4.24-4.45 (m, 5H),4.08-4.21 (m, 1H), 3.85 (t, J=9.70 Hz, 1H), 3.60-3.79 (m, 3H), 3.32-3.46(m, 2H), 3.24 (d, J=9.70 Hz, 1H), 3.15 (d, J=9.70 Hz, 1H), 2.59 (dd,J=14.55, 4.41 Hz, 1H), 2.32-2.52 (m, 2H), 2.17-2.31 (m, 2H), 1.98-2.15(m, 1H), 1.78-1.97 (m, 3H), 1.37-1.78 (m, 8H), 1.33 (d, J=6.17 Hz, 3H),1.18-1.27 (m, 3H), 1.12 (d, J=6.17 Hz, 3H), 1.05 (d, J=7.06 Hz, 3H).LCMS (ESI): m/z: [M+Na] calcd for C₅₄H₈₀ClN₃O₁₆Na: 1084.52; found:1084.5.

Example 20. Synthesis of Compound 2-BS

Compound 2-BS was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with1-methylazetidin-3-amine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm6.00-6.60 (m, 13H) 5.51 (d, J=5.29 Hz, 1H) 5.32-5.42 (m, 2H) 4.82 (s,1H) 4.39-4.71 (m, 4H) 4.22-4.37 (m, 2H) 3.78-4.19 (m, 5H) 3.40-3.76 (m,7H) 3.21-3.33 (m, 6H) 2.53 (s, 3H) 2.32-2.46 (m, 2H) 2.19-2.26 (m, 1H)1.35-2.16 (m, 13H) 1.33 (d, J=6.17 Hz, 3H) 1.23 (d, J=6.62 Hz, 3H)1.08-1.16 (m, 3H) 1.04 (d, J=7.06 Hz, 3 H). LCMS (ESI): m/z: [M+H] calcdfor C₅₁H₈₃N₄O₁₆: 1007.57; found 1007.5.

Example 21. Synthesis of Compound 2-Z

Compound 2-Z was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-(2-methoxyethoxy)ethanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δppm 8.75-8.73 (1H, M) 6.54-6.19 (14H, m), 5.49-5.39 (2H, m), 4.80-3.28(25H, m), 2.23-0.92 (30H, m). LCMS (ESI): m/z: [M+Na] calcd forC₅₂H₈₅N₃O₁₈: 1062.6; found 1062.6.

Example 22. Synthesis of Compound 2-AE

Compound 2-AE was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(R)-1-(2-aminoethyl)pyrrolidin-3-ol. ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 6.48-6.15 (14H, m), 5.37-5.32 (2H, m),4.83-2.86 (22H, m), 2.25-0.97 (33H, m). LCMS (ESI): m/z: [M+H] calcd forC₅₃H₈₇N₄O₁₇: 1051.6; found 1051.6.

Example 23. Synthesis of Compound 2-AH

Compound 2-AH was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with phenylmethanamine.¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 8.73 (1H, m), 7.31-7.12 (5H,m), 6.49-6.16 (14H, m), 5.37-5.27 (2H, m), 4.79-3.23 (16H, m) 2.36-1.04(30H, m). LCMS (ESI): m/z: [M+Na] calcd for C₅₄H₈₁N₃O₁₆Na: 1050.6; found1050.6.

Example 24. Synthesis of Compound 2-E

Compound 2-E was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with morpholine. ¹H NMR(400 MHz, Methanol-d4+Pyr-d5): δ 6.09-6.57 (m, 13H) 5.51 (d, J=6.17 Hz,1H) 4.80 (s, 1H) 4.64 (br. S., 1H) 4.52 (br. S., 1H) 4.31-4.45 (m, 2H)4.20-4.31 (m, 2H) 3.62-3.90 (m, 5H) 3.52 (d, J=4.41 Hz, 4H) 3.39 (d,J=4.41 Hz, 5H) 3.18-3.27 (m, 2H) 2.32-2.53 (m, 3H) 2.17-2.31 (m, 2H)2.04 (d, J=11.03 Hz, 2H) 1.87 (d, J=7.94 Hz, 3H) 1.62-1.79 (m, 3H)1.27-1.62 (m, 10H) 1.23 (d, J=6.17 Hz, 3H) 1.12 (d, J=6.17 Hz, 3H) 1.04(d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd for C₅₁H₅₁N₃O₁₇Na:1030.6; found 1030.6.

Example 25. Synthesis of Compound 2-AG

Compound 2-AG was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with cyclobutanamine. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ 6.05-6.63 (m, 13H) 4.82 (s, 1H)4.66 (br. s., 1H) 4.52 (t, J=10.54 Hz, 1H) 4.18-4.43 (m, 4H) 4.03-4.14(m, 1H) 3.85 (br. s., 1H) 3.64-3.78 (m, 2H) 3.60 (br. s., 1H) 3.46 (dd,J=8.78, 6.27 Hz, 1H) 3.27 (d, J=9.54 Hz, 1H) 1.96-2.57 (m, 10H)1.62-1.96 (m, 9H) 1.38-1.63 (m, 8H) 1.35 (d, J=6.02 Hz, 4H) 1.25 (d,J=6.53 Hz, 3H) 1.15 (d, J=6.53 Hz, 3H) 1.07 (d, J=7.53 Hz, 3H). LCMS(ESI): m/z: [M+Na] calcd for C₅₁H₁N₃O₁₆Na: 1014.5; found 1014.5.

Example 26. Synthesis of Compound 2-AI

Compound 2-AI was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with pyrrolidine. ¹H NMR(400 MHz, Methanol-d4+Pyr-d5): 66.10-6.61 (m, 12H) 5.53 (d, J=5.52 Hz,1H) 4.83 (s, 1H) 4.68 (br. s., 1H) 4.51-4.61 (m, 1H) 4.24-4.48 (m, 4H)3.67-3.92 (m, 4H) 3.19-3.31 (m, 5H) 2.20-2.56 (m, 5H) 2.00-2.14 (m, 1H)1.84-2.00 (m, 3H) 1.30-1.83 (m, 17H) 1.26 (d, J=6.02 Hz, 3H) 1.14 (d,J=6.02 Hz, 3H) 1.07 (d, J=7.53 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcdfor C₅₁H₁N₃O₁₆Na: 1014.5; found 1014.5.

Example 27. Synthesis of Compound 2-BT

Compound 2-BT was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with ethanamine. ¹H NMR(400 MHz, Methanol-d4+Pyr-d5): δ 8.69 (s, 1H), 6.14-6.48 (m, 13H),5.36-5.49 (m, 3H), 4.84 (s, 1H), 4.81 (br. s., 1H), 4.64 (t, 1H),4.48-4.53 (m, 1H), 4.24-4.34 (m, 3H), 4.06-4.08 (m, 1H), 3.74-3.84 (m,1H), 3.60-3.74 (m, 1H), 3.42-3.47 (m, 1H), 3.23-3.26 (m, 2H), 3.13-3.17(m, 3H), 2.34-2.50 (m, 3H), 2.20-2.24 (m, 2H), 1.86 (m, 2H), 1.41-1.72(m, 7H), 1.31 (d, J=6 Hz, 3H), 1.23 (d, J=6 Hz, 3H), 1.11 (d, J=6 Hz,3H), 0.98 (d, J=6 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd forC₄₉H₇₉N₃O₁₆Na: 988.55; found 988.5.

Example 28. Synthesis of Compound 2-BU

Compound 2-BU was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with cyclopropanamine. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 7.37 (d, J=8.03 Hz, 2H) 7.21(t, J=7.78 Hz, 2H) 6.97 (t, J=7.28 Hz, 1H) 6.09-6.57 (m, 13H) 5.50 (br.s., 2H) 5.36-5.45 (m, 2H) 4.77 (s, 1H) 4.63 (br. s., 1H) 4.46-4.55 (m,2H) 4.27-4.39 (m, 3H) 4.21 (d, J=3.01 Hz, 1H) 4.04-4.15 (m, 2H)3.79-3.89 (m, 2H) 3.73 (d, J=11.04 Hz, 2H) 3.53-3.66 (m, 3H) 3.42 (dd,J=9.03, 6.02 Hz, 1H) 3.26 (br. s., 1H) 3.12 (d, J=9.03 Hz, 3H) 2.50-2.59(m, 2H) 2.41-2.49 (m, 2H) 2.33-2.40 (m, 1H) 2.19-2.29 (m, 3H) 1.95-2.06(m, 2H) 1.81-1.94 (m, 4H) 1.37-1.79 (m, 12H) 1.33 (d, J=6.02 Hz, 3H)1.25 (d, J=6.53 Hz, 3H) 1.15 (d, J=6.53 Hz, 3H) 1.07 (d, J=7.53 Hz, 3H)0.60 (d, J=7.03 Hz, 3H) 0.46 (br. s., 3H). LCMS (ESI): m/z: [M+Na] calcdfor C₅₀H₇₉N₃O₁₆Na: 1000.55; found 1000.5.

Example 29. Synthesis of Compound 2-BV

Compound 2-BV was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with3,3-difluorocyclobutanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm6.03-6.56 (m, 13H) 5.48 (d, J=6.62 Hz, 1H) 5.31-5.39 (m, 2H) 4.72 (s,1H) 4.60 (br. s., 1H) 4.48 (t, J=10.14 Hz, 1H) 4.20-4.37 (m, 2H)4.10-4.19 (m, 2H) 3.97-4.09 (m, 1H) 3.81 (t, J=9.48 Hz, 1H) 3.70 (d,J=10.58 Hz, 1H) 3.44-3.63 (m, 2H) 3.31-3.41 (m, 1H) 3.22 (d, J=9.26 Hz,1H) 2.66-3.02 (m, 3H) 2.28-2.63 (m, 5H) 2.13-2.26 (m, 2H) 1.98 (d,J=8.38 Hz, 1H) 1.59-1.91 (m, 5H) 1.32-1.58 (m, 6H) 1.30 (d, J=6.17 Hz,3H) 1.21 (d, J=6.62 Hz, 3H) 1.10 (d, J=6.62 Hz, 3H) 1.02 (d, J=7.06 Hz,3H). LCMS (ESI): m/z: [M−H₂O] calcd for C₅₁H₇₉F₂N₃O₁₆: 1027.54; found1010.3.

Example 30. Synthesis of Compound 2-BW

Compound 2-BW was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(9H-fluoren-9-yl)methyl 2-(aminomethyl)pyrrolidine-1-carboxylate. ¹H NMR(400 MHz, Methanol-d4+Pyr-d5): δ 8.73 (s, 2H) 6.46-6.49 (m, 2H)6.18-6.33 (m, 11H) 5.5 (s, 2H) 4.75 (s, 1H) 4.52-4.61 (m, 1H) 4.44-4.52(m, 2H) 4.24-4.34 (m, 3H) 4.12 (s, 1H) 3.72-3.83 (m, 2H) 3.57 (t, 1H)3.51 (s, 2H) 3.04-3.10 (m, 3H) 2.19-2.42 (m, 3H) 1.53-1.87 (m, 16H) 1.28(d, J=6 Hz, 3H) 1.23 (d, J=6 Hz, 3H) 1.11 (d, J=6 Hz, 3H) 1.04 (d, J=6Hz, 3H). LCMS (ESI): m/z: [M+H] calcd for C₅₂H₈₄N₄O₁₆: 1021.59; found1021.5.

Example 31. Synthesis of Compound 2-G

Compound 2-G was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(1S,3S)-cyclobutane-1,3-diamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ8.76 (s, 1H) 6.16-6.56 (m, 13H) 5.54-5.55 (m, 1H) 4.86 (s, 1H) 4.46-4.56(m, 2H) 4.38-(m, 2H) 4.12-4.18 (m, 2H) 3.68-3.88 (m, 3H) 3.38-3.49 (m,2H) 3.38-3.15 (m, 1H) 3.22-3.24 (m, 1H) 1.38-2.74 (m, 22H) 1.32 (d, J=6Hz, 3H) 1.24 (d, J=6 Hz, 3H) 1.11 (d, J=6 Hz, 3H) 1.04 (d, J=6 Hz, 3H).LCMS (ESI): m/z: [M+H] calcd for C₅₁H₈₂N₄O₁₆: 1007.57; found 1007.5.

Example 32. Synthesis of Compound 2-BX

Compound 2-BX was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withtrans-cyclobutane-1,3-diamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δppm 8.67-8.75 (m, 1H) 6.07-6.62 (m, 13H) 5.52 (d, J=6.17 Hz, 1 H)5.34-5.44 (m, 2H) 4.83 (s, 1H) 4.23-4.73 (m, 5H) 4.12 (td, J=10.36, 4.85Hz, 1H) 3.59-3.95 (m, 5H) 3.32-3.51 (m, 3H) 3.24 (d, J=9.26 Hz, 1H)2.18-2.63 (m, 8H) 1.95-2.12 (m, 1H) 1.61-1.94 (m, 6H) 1.33-1.61 (m, 6H)1.31 (d, J=5.73 Hz, 3H) 1.23 (d, J=6.17 Hz, 3H) 1.12 (d, J=6.17 Hz, 3H)1.04 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd forC₅₁H₈₂N₄O₁₆Na: 1029.57; found 1029.5.

Example 33. Synthesis of Compound 2-L

Compound 2-L was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-methyl-2,6-diazaspiro[3.3]heptane. ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 6.29 (br. s., 13H), 5.50 (d, J=4.41 Hz, 2H),4.83 (s, 1H), 4.60-4.69 (m, 1H), 4.46-4.57 (m, 1H), 4.34 (d, J=3.09 Hz,3H), 4.23 (d, J=3.97 Hz, 2H), 3.95-4.06 (m, 4H), 3.62-3.88 (m, 5H),3.39-3.57 (m, 4H), 3.14 (d, J=8.82 Hz, 1H), 2.91 (d, J=14.55 Hz, 1H),2.31-2.51 (m, 4H), 2.14-2.30 (m, 5H), 1.27-2.09 (m, 19H), 1.23 (d,J=6.17 Hz, 3H), 1.12 (d, J=6.17 Hz, 3H), 1.00-1.09 (m, 4H). LCMS (ESI):m/z: [M+Na] calcd for C₅₃H₈₄N₄O₁₆Na: 1055.6; found 1055.6.

Example 34. Synthesis of Compound 2-BY

Compound 2-BY was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withN₁-methyl-N₁-(oxetan-3-yl)ethane-1,2-diamine. ¹H NMR (500 MHz,Pyridine-d5: Methanol-d₄=1:1): δ 8.76 (d, J=104.4 Hz, 2H), 7.76 (d,J=7.5 Hz, 1H), 7.32 (t, J=7.5 Hz, 1H), 6.69-5.78 (m, 7H), 5.51 (d, J=6.5Hz, 1H), 5.37 (dd, J=14.7, 10.2 Hz, 1H), 4.79 (d, J=7.5 Hz, 1H), 4.64(s, 1H), 4.58-4.44 (m, 4H), 4.39-4.30 (m, 1H), 4.25 (d, J=20.8 Hz, 2H),4.07 (d, J=11.9 Hz, 1H), 3.89-3.67 (m, 2H), 3.69-3.55 (m, 2H), 3.51-3.35(m, 2H), 3.26 (t, J=7.3 Hz, 3H), 2.79 (t, J=11.9 Hz, 1H), 2.52 (d,J=15.8 Hz, 1H), 2.47-2.35 (m, 2H), 2.34 (d, J=4.7 Hz, 1H), 2.28-2.16 (m,4H), 1.99 (s, 2H), 1.85 (q, J=15.1, 11.1 Hz, 2H), 1.77-1.62 (m, 1H),1.55 (dd, J=13.2, 8.4 Hz, 2H), 1.47 (s, 1H), 1.42-1.37 (m, 1H), 1.33 (d,J=6.4 Hz, 3H), 1.23 (d, J=6.3 Hz, 3H), 1.12 (d, J=6.4 Hz, 3H), 1.05 (d,J=7.1 Hz, 3H), 0.76 (s, 1H). LCMS (ESI): Calcd for C₅₃H₈₆N₄O₁₇: 1051.28;m/Z: [M+H] found 1052.40.

Example 35. Synthesis of Compound 2-BZ

Compound 2-BZ was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(S)-1-methylpyrrolidin-3-amine. ¹H NMR (500 MHz, Pyridine-d5:Methanol-d₄=1:1): δ 8.77 (s, 1H), 6.55-6.34 (m, 2H), 6.34-6.26 (m, 9H),6.19 (ddt, J=19.9, 14.0, 6.5 Hz, 1H), 5.52 (d, J=7.3 Hz, 1H), 5.38 (dd,J=14.6, 10.2 Hz, 1H), 4.80 (d, J=16.4 Hz, 1H), 4.65 (s, 1H), 4.52 (t,J=10.8 Hz, 1H), 4.41-4.18 (m, 2H), 4.07 (d, J=12.0 Hz, 1H), 3.88-3.69(m, 2H), 3.69-3.41 (m, 1H), 3.25 (d, J=9.5 Hz, 1H), 2.72 (d, J=26.5 Hz,2H), 2.54-2.34 (m, 2H), 2.29 (s, 2H), 2.27-2.19 (m, 2H), 1.89 (d, J=12.6Hz, 2H), 1.55 (t, J=12.0 Hz, 2H), 1.33 (dd, J=9.4, 6.1 Hz, 3H), 1.24 (d,J=6.3 Hz, 3H), 1.13 (d, J=6.4 Hz, 3H), 1.05 (d, J=7.0 Hz, 3H). LCMS(ESI): Calcd for C₅₂H₈₄N₄O₁₆: 1020.59; m/Z: [M+H] found 1021.35.

Example 36. Synthesis of Compound 2-CA

Compound 2-CA was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(R)-1-methylpyrrolidin-3-amine. ¹H NMR (500 MHz, Methanol-d₄): δ 8.52(s, 2H), 6.53-6.06 (m, 11H), 5.95 (dd, J=15.4, 8.8 Hz, 1H), 5.38 (dd,J=13.7, 9.0 Hz, 2H), 4.58 (s, 1H), 4.45 (d, J=8.5 Hz, OH), 4.33 (dd,J=24.7, 14.1 Hz, 1H), 4.18 (t, J=9.8 Hz, 1H), 4.11-3.87 (m, 1H),3.86-3.64 (m, 2H), 3.65-3.40 (m, 1H), 3.42-3.31 (m, 2H), 3.26-3.06 (m,2H), 3.07-2.96 (m, 1H), 2.95-2.72 (m, 1H), 2.67 (d, J=11.1 Hz, 3H), 2.40(q, J=7.5, 6.8 Hz, 2H), 2.29 (dd, J=17.2, 9.8 Hz, 1H), 2.19 (dd, J=17.0,2.6 Hz, 1H), 2.15-1.77 (m, 1H), 1.72 (dd, J=13.6, 8.3 Hz, 3H), 1.59 (d,J=13.9 Hz, 1H), 1.53-1.29 (m, 3H), 1.28 (d, J=6.1 Hz, 4H), 1.19 (d,J=6.4 Hz, 3H), 1.11 (d, J=6.4 Hz, 3H), 1.01 (d, J=7.2 Hz, 3H). LCMS(ESI): Calcd for C₅₂H₈₄N₄O₁₆: 1020.59; m/Z: [M+H]: found 1022.35.

Example 37. Synthesis of Compound 2-CB

Compound 2-CB was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-(2-oxa-6-azaspiro[3.3]heptan-6-yl)ethan-1-amine. ¹H NMR (500 MHz,Methanol-d₄): δ 8.55 (s, 2H), 6.31 (dddd, J=62.3, 48.4, 19.6, 8.8 Hz,10H), 5.94 (dd, J=15.2, 9.0 Hz, 1H), 5.48-5.29 (m, 2H), 4.76 (s, 4H),4.58 (s, 1H), 4.18 (t, J=9.8 Hz, 1H), 4.10-3.91 (m, 2H), 3.82 (td,J=10.5, 4.7 Hz, 1H), 3.74 (s, 4H), 3.60 (d, J=11.0 Hz, 1H), 3.38 (t,J=9.6 Hz, 1H), 3.29-3.08 (m, 2H), 3.10-2.97 (m, 1H), 2.75 (d, J=7.5 Hz,2H), 2.38 (d, J=6.8 Hz, OH), 2.29 (dd, J=17.2, 9.8 Hz, 1H), 2.25-2.13(m, 2H), 2.13-1.94 (m, 1H), 1.86-1.66 (m, 4H), 1.59 (d, J=13.8 Hz, 1H),1.28 (d, J=6.3 Hz, 3H), 1.20 (d, J=6.4 Hz, 3H), 1.12 (d, J=6.4 Hz, 3H),1.02 (d, J=7.2 Hz, 3H). LCMS (ESI): Calcd for: C₅₄H₈₆N₄O₁₇ 1062.60; m/Z:[M+H] found 1063.50.

Example 38. Synthesis of Compound 2-CC

Compound 2-CC was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(R)-1-methyl-2-aminomethylpyrrolidine. ¹H NMR (500 MHz, Methanol-d₄): δ8.53 (s, 4H), 6.51-6.13 (m, 8H), 5.99 (dd, J=15.3, 9.1 Hz, 1H),5.42-5.33 (m, 2H), 4.55 (s, 1H), 4.45-4.32 (m, 2H), 4.25 (s, 1H), 4.18(s, 2H), 4.03-3.94 (m, 1H), 3.91 (d, J=3.1 Hz, 1H), 3.72 (t, J=9.2 Hz,1H), 3.61 (d, J=10.8 Hz, 1H), 3.52-3.44 (m, 2H), 3.36 (d, J=8.2 Hz, 1H),3.23-3.15 (m, 2H), 3.09-3.00 (m, 2H), 2.84 (d, J=9.7 Hz, 1H), 2.70 (s,3H), 2.38 (d, J=5.6 Hz, 1H), 2.30 (dd, J=17.0, 9.8 Hz, 1H), 2.23-2.08(m, 4H), 2.08-1.97 (m, 2H), 1.84-1.64 (m, 6H), 1.61 (d, J=14.1 Hz, 1H),1.53-1.29 (m, 5H), 1.28 (d, J=6.3 Hz, 3H), 1.20 (d, J=6.3 Hz, 3H), 1.12(d, J=6.3 Hz, 3H), 1.02 (d, J=7.2 Hz, 3H). LC-MS: Calculated(C₅₃H₈₆N₄O₁₆+H)⁺: 1035.61. Observed: 1036.45.

Example 39. Synthesis of Compound 2-CD

Compound 2-CD was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(S)-1-methyl-2-aminomethylpyrrolidine. ¹H NMR (500 MHz, Methanol-d₄): δ8.53 (s, 4H), 6.50-6.12 (m, 8H), 5.99 (dd, J=14.9, 9.1 Hz, 1H),5.42-5.34 (m, 2H), 4.58 (s, 1H), 4.46-4.31 (m, 2H), 4.18 (s, 3H), 3.96(d, J=16.6 Hz, 2H), 3.76-3.67 (m, 1H), 3.60 (d, J=11.3 Hz, 1H),3.47-3.37 (m, 2H), 3.28-3.13 (m, 3H), 3.09-3.01 (m, 2H), 2.90 (t, J=12.5Hz, 1H), 2.71 (d, J=8.6 Hz, 3H), 2.38 (d, J=7.0 Hz, 1H), 2.29 (dd,J=17.2, 9.8 Hz, 1H), 2.25-2.07 (m, 4H), 2.04-1.96 (m, 2H), 1.83-1.68 (m,6H), 1.60 (d, J=14.0 Hz, 1H), 1.52-1.32 (m, 5H), 1.28 (d, J=6.2 Hz, 3H),1.20 (d, J=6.5 Hz, 3H), 1.12 (d, J=6.4 Hz, 3H), 1.02 (d, J=7.2 Hz, 3H).LC-MS: Calculated (C₅₃H₈₆N₄O₁₆+H)⁺: 1035.61. Observed: 1036.45.

Example 40. Synthesis of Compound 2-CE Methyl Ketal

Compound 2-CE methyl ketal was synthesized in the manner similar toCompound 2-3-I, except piperazine was substituted with methyl amine, andformic acid was not used in the mobile phase (acetonitrile/water) duringpurification. ¹H NMR (500 MHz, Methanol-d₄): δ 6.49-6.10 (m, 12H), 5.86(dd, J=14.3, 7.1 Hz, 1H), 5.45 (dd, J=14.0, 9.5 Hz, 1H), 5.30-5.18 (m,1H), 4.61 (t, J=7.3 Hz, 1H), 4.52 (s, 1H), 4.20-4.11 (m, 1H), 3.99-3.91(m, 1H), 3.80 (s, 1H), 3.71 (dd, J=9.5, 5.0 Hz, 2H), 3.66 (t, J=9.0 Hz,1H), 3.51 (d, J=10.5 Hz, 1H), 3.41-3.34 (m, 2H), 3.30-3.04 (m, 7H), 2.72(s, 3H), 2.53 (s, 1H), 2.42-2.34 (m, 1H), 2.34-2.22 (m, 2H), 2.17 (dd,J=14.8, 7.3 Hz, 1H), 1.92-1.78 (m, 2H), 1.78-1.65 (m, 3H), 1.64-1.57 (m,2H), 1.52 (t, J=12.2 Hz, 1H), 1.44 (tdd, J=14.0, 12.7, 10.4, 5.8 Hz,5H), 1.27 (d, J=5.9 Hz, 3H), 1.20 (d, J=6.4 Hz, 3H), 1.11 (d, J=6.5 Hz,3H), 1.01 (d, J=7.2 Hz, 3H). LC-MS: Calculated (C₄₉H₇₉N₃O₁₆+H)⁺: 966.55.Observed: 966.50.

Example 41. Synthesis of Compound 2-CF

Compound 2-CF was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-methyl-2-morpholinopropan-1-amine. ¹H NMR (500 MHz, Methanol-d₄): δ7.84 (d, J=7.5 Hz, 3H), 7.78 (d, J=7.5 Hz, 1H), 7.73 (d, J=7.6 Hz, 1H),7.66 (d, J=7.4 Hz, 3H), 7.42 (t, J=7.4 Hz, 3H), 7.36 (td, J=7.4, 1.2 Hz,4H), 7.33-7.28 (m, 1H), 6.55-6.13 (m, 15H), 5.93 (dd, J=15.1, 9.0 Hz,1H), 5.36 (d, J=8.5 Hz, 3H), 4.57 (d, J=10.3 Hz, 3H), 4.45 (d, J=8.1 Hz,1H), 4.34 (t, J=10.4 Hz, 1H), 4.22-4.13 (m, 3H), 4.03 (d, J=8.9 Hz, 1H),3.92 (d, J=3.1 Hz, 1H), 3.84-3.75 (m, 1H), 3.75-3.53 (m, 16H), 3.44 (d,J=5.2 Hz, 7H), 3.35 (s, 2H), 3.29-3.21 (m, 4H), 3.23-3.07 (m, 8H), 3.01(s, 2H), 2.89 (s, 2H), 2.66 (s, 45H), 2.59 (s, 13H), 2.60-2.53 (m, 3H),2.52 (d, J=15.4 Hz, 4H), 2.36 (t, J=4.7 Hz, 7H), 2.24-2.14 (m, 2H), 2.03(s, 4H), 1.83-1.72 (m, 2H), 1.72 (s, 2H), 1.59 (d, J=13.6 Hz, 3H),1.53-1.37 (m, 3H), 1.36-1.24 (m, 7H), 1.20 (d, J=6.4 Hz, 5H), 1.12 (d,J=7.9 Hz, 8H), 1.07-0.97 (m, 31H). LC-MS: Calculated (C₅₅H₉₀N₄O₁₇+H)⁺:1080.34. Observed: 1080.40.

Example 42. Synthesis of Compound 2-CG

Compound 2-CG was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withspiro[3.3]heptan-2-amine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ 8.76(s, 1H) 6.04-6.62 (m, 13H) 5.51 (d, J=5.73 Hz, 1H) 4.80 (s, 1H) 4.64(br. s., 1H) 4.51 (t, J=10.14 Hz, 1H) 4.20-4.38 (m, 3H) 4.01-4.18 (m,2H) 3.84 (br. s., 1H) 3.55-3.76 (m, 3H) 3.39-3.50 (m, 1H) 3.24 (d,J=9.70 Hz, 2H) 2.28-2.58 (m, 4H) 2.14-2.25 (m, 3H) 1.96-2.08 (m, 1H)1.37-1.93 (m, 23H) 1.33 (d, J=6.17 Hz, 4H) 1.23 (d, J=6.17 Hz, 3H) 1.12(d, J=6.17 Hz, 3H) 1.04 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+H] calcdfor C₅₅H₈₇N₃O₁₆: 1032.59; found 1032.6.

Example 43. Synthesis of Compound 2-CH

Compound 2-CH was synthesized in the manner similar to Compound 2-I(Example 3) and 2-CL (Example 55), except piperazine was substitutedwith allyl (2-aminopropyl)carbamate (3). ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 6.11-6.61 (m, 10H), 5.55 (d, J=6.02 Hz, 1H),4.80-4.90 (m, 1H), 4.60-4.75 (m, 1H), 4.56 (d, J=18.70 Hz, 1H), 4.45(br. s., 1H), 4.24-4.41 (m, 2H), 4.15 (d, J=6.27 Hz, 1H), 3.58-3.90 (m,4H), 3.42-3.56 (m, 3H), 3.12-3.22 (m, 2H), 2.98-3.08 (m, 1H), 2.90 (dd,J=14.62, 2.95 Hz, 1H), 2.71-2.83 (m, 1H), 2.33-2.68 (m, 3H), 2.17-2.31(m, 2H), 1.31-2.09 (m, 20H), 1.20-1.31 (m, 6H), 1.15 (d, J=6.27 Hz, 3H),1.02-1.10 (m, 5H), 0.72 (s, 2H). LCMS (ESI): m/z: [M+Na] calcd forC₅₀H₈₂N₄O₁₆Na: 1017.5; found 1017.5.

Synthesis of allyl (2-aminopropyl)carbamate (3)

Step 1: To a solution of compound 1 (800.00 mg, 4.59 mmol, 1.00 equiv.)in DCM (6.00 mL) was added allyl carbonochloridate (1.66 g, 13.77 mmol,1.46 mL, 3.00 equiv.) slowly at 0° C. The mixture was stirred at 25° C.for 1 hour. The mixture was quenched by addition 10% citric acidsolution, and extracted with DCM (45 mL). The combined organic layerswere dried over Na₂SO₄, filtered and concentrated under reduced pressureto give the 1.4 g of compound 2. ¹H NMR (400 MHz, CDCl₃-d): δ ppm 1.07(d, J=6.62 Hz, 3H), 1.27-1.42 (m, 10H), 2.94-3.28 (m, 3H), 3.43 (d,J=7.50 Hz, 1H), 3.70 (br. s., 1H), 4.39-4.63 (m, 2H), 5.04-5.31 (m, 2H), 5.78-5.95 (m, 1H).

Step 2: To a solution of compound 2 (1.40 g, 5.42 mmol, 1.00 equiv.) inMeOH (5.00 mL) was added HCl/MeOH (1 M, 5.42 mL, 1.00 equiv.). Themixture was stirred at 25° C. for 1 hour, and concentrated under reducedpressure to give 1.2 g of compound 3. The residue was neutralized byIon-exchange resin. ¹H NMR (400 MHz, Methanol-d4): δ ppm 1.26 (d, J=6.62Hz, 4H), 3.26-3.31 (m, 5H), 4.55 (d, J=4.85 Hz, 2H), 5.18 (d, J=10.14Hz, 1H), 5.30 (d, J=17.20 Hz, 1H), 5.85-6.03 (m, 1H).

Example 44. Synthesis of Compound 2-CI

Compound 2-CI was synthesized in the manner similar to Compound 2-I(Example 3) and 2-CL (Example 55), except piperazine was substitutedwith allyl(1-aminopropan-2-yl)carbamate (3). ¹H NMR (400 MHz,Methanol-d4+Py-d5): δ ppm 6.07-6.64 (m, 9H), 5.53 (d, J=6.17 Hz, 1H),4.85 (br. s., 1H), 4.67 (br. s., 1H), 4.45-4.59 (m, 2H), 4.33 (d, J=7.50Hz, 2H), 4.09-4.23 (m, 1H), 3.39-4.03 (m, 10H), 2.67-2.93 (m, 1H),2.32-2.67 (m, 4H), 1.96-2.31 (m, 4H), 1.40-1.95 (m, 12H), 1.18-1.40 (m,8H), 1.12 (d, J=6.17 Hz, 3H), 0.98-1.07 (m, 3H). LCMS (ESI): m/z: [M+Na]calcd for C₅₀H₈₂N₄O₁₆Na: 1017.6; found 1017.6.

Synthesis of allyl(1-aminopropan-2-yl)carbamate (3)

Step 1: To a solution of allyl carbonochloridate (1.66 g, 13.77 mmol,1.46 mL, 3.00 equiv.) in DCM (10.00 mL) was added compound 1 (800.00 mg,4.59 mmol, 1.00 equiv.) slowly at 0° C. The mixture was stirred at 25°C. for 1 hour. The mixture was quenched by addition 10% citric acidsolution, and extracted with DCM (45 mL). The combined organic layerswere dried over Na₂SO₄, and filtered. Concentratration under reducedpressure resulted in 1.2 g of compound 2.

Step 2: To a solution of compound 2 (500.00 mg, 1.94 mmol, 1.00 equiv.)in MeOH (2.00 mL) was added HCl/MeOH (1 M, 1.94 mL, 1.00 equiv.). Themixture was stirred at 25° C. for 1 hour. The mixture was concentratedunder reduced pressure to give 380 mg of compound 3. The residue wasalkalized by Ion-exchange resin and submitted to the next step withoutfurther purification.

Example 45. Synthesis of Compound 2-AD

Compound 2-AD was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with4-(azetidin-3-yl)morpholine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm6.05-6.62 (m, 11H), 4.78-4.91 (m, 1H), 4.47-4.72 (m, 1H), 4.11-4.46 (m,4H), 3.66-4.09 (m, 5H), 3.55 (br. s., 4H), 2.78-3.01 (m, 1H), 2.50 (s,1H), 1.99-2.33 (m, 4H), 1.89 (br. s., 2H), 1.65-1.83 (m, 1H), 1.43-1.64(m, 1H), 1.33 (d, J=5.73 Hz, 3H), 1.23 (br. s., 2H), 1.12 (d, J=6.17 Hz,3H), 1.01-1.07 (m, 1H). LCMS (ESI): m/z: [M+Na] calcd for C₅₄H₈₆N₄O₁₇Na:1085.6; found 1085.6.

Example 46. Synthesis of Compound 2-CM

Compound 2-CM was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withcyclobutylmethanamine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm6.08-6.56 (m, 12H), 5.45-5.57 (m, 1H), 5.34-5.42 (m, 1H), 4.75-4.83 (m,1H), 4.60-4.68 (m, 1H), 4.45-4.55 (m, 1H), 4.30-4.40 (m, 1H), 4.26 (br.s., 2H), 4.00-4.11 (m, 1H), 3.84 (br. s., 1H), 3.73 (d, J=11.03 Hz, 1H),3.61 (t, J=9.26 Hz, 2H), 3.43 (br. s., 1H), 3.20-3.27 (m, 1H), 3.07-3.20(m, 3H), 2.29-2.58 (m, 4H), 2.16-2.27 (m, 2H), 1.96-2.08 (m, 1H),1.77-1.94 (m, 6H), 1.62-1.76 (m, 5H), 1.49-1.62 (m, 5H), 1.44-1.49 (m, 1H), 1.39-1.43 (m, 1H), 1.32 (d, J=5.73 Hz, 4H), 1.23 (d, J=6.17 Hz, 3H),0.93-1.16 (m, 6H). LCMS (ESI): m/z: [M+H] calcd for C₅₂H₈₄N₃O₁₆: 1006.5;found 1006.5.

Example 47. Synthesis of Compound 2-CN

Compound 2-CN was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with isopropylamine. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.05-6.57 (m, 14H), 5.44-5.56(m, 1H), 5.32-5.43 (m, 2H), 4.76 (s, 1H), 4.57-4.67 (m, 1H), 4.43-4.54(m, 1H), 4.27-4.38 (m, 1H), 4.18 (d, J=2.65 Hz, 2H), 3.96-4.08 (m, 1H),3.86 (dt, J=13.01, 6.73 Hz, 3H), 3.67-3.75 (m, 1H), 3.48-3.63 (m, 2H),3.40 (br. s., 1H), 3.24 (d, J=9.70 Hz, 1H), 2.99-3.10 (m, 1H), 2.31-2.51(m, 3H), 2.17-2.26 (m, 2H), 1.93-2.04 (m, 1H), 1.61-1.92 (m, 7H),1.54-1.59 (m, 1H), 1.51-1.54 (m, 1H), 1.48-1.51 (m, 1H), 1.43-1.48 (m,1H), 1.39-1.43 (m, 1H), 1.36-1.39 (m, 1H), 1.33-1.36 (m, 1H), 1.31 (d,J=6.17 Hz, 3H), 1.23 (d, J=6.17 Hz, 3H), 1.13 (br. s., 2H), 1.11 (br.s., 2H), 1.10 (s, 1 H), 1.08 (s, 2H), 1.05 (br. s., 2H), 1.03 (s, 4H),1.01 (br. s., 3H). LCMS (ESI): m/z: [M+Na] calcd for C₅₀H₅₁N₃O₁₆Na:1002.5; found 1002.5.

Example 48. Synthesis of Compound 2-CO

Compound 2-CO was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with isobutylamine. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.06-6.57 (m, 15H), 5.44-5.56(m, 1H), 5.33-5.42 (m, 2H), 4.77 (s, 1H), 4.57-4.68 (m, 1H), 4.44-4.54(m, 1H), 4.33 (br. s., 1H), 4.22 (br. s., 2H), 3.99-4.09 (m, 1H), 3.83(br. s., 1H), 3.72 (d, J=11.03 Hz, 1H), 3.57 (t, J=9.48 Hz, 2H),3.36-3.46 (m, 1H), 3.24 (d, J=9.26 Hz, 1H), 3.09 (d, J=9.26 Hz, 1H),2.87-3.03 (m, 3H), 2.34 (d, J=9.70 Hz, 3H), 2.16-2.28 (m, 2H), 2.01 (br.s., 1H), 1.77-1.92 (m, 3H), 1.60-1.77 (m, 4H), 1.55-1.60 (m, 1H),1.52-1.55 (m, 2H), 1.49-1.52 (m, 1H), 1.44-1.49 (m, 1H), 1.39-1.43 (m,1H), 1.36-1.39 (m, 1H), 1.33-1.36 (m, 1H), 1.31 (d, J=5.73 Hz, 3H), 1.23(d, J=6.62 Hz, 3H), 1.12 (d, J=6.17 Hz, 3H), 1.04 (d, J=7.06 Hz, 3H).LCMS (ESI): m/z: [M+Na] calcd for C₅₁H₈₃N₃O₁₆Na: 1016.5; found 1016.5.

Example 49. Synthesis of Compound 2-CP

Compound 2-CP was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with cyclohexylamine. ¹HNMR (400 MHz, Methanol-d4+Py-d5): δ ppm 6.08-6.63 (m, 13H), 5.53 (d,J=5.29 Hz, 2H), 4.74 (s, 1H), 4.64 (br. s., 1H), 4.52 (t, J=9.92 Hz,1H), 4.21-4.43 (m, 3H), 4.00-4.14 (m, 3H), 3.84 (t, J=9.26 Hz, 2H), 3.73(d, J=11.03 Hz, 1H), 3.52-3.69 (m, 3H), 3.37-3.45 (m, 2H), 3.23 (d,J=9.70 Hz, 2H), 2.76 (d, J=7.50 Hz, 1H), 2.53-2.62 (m, 1H), 2.32-2.50(m, 3H), 2.16-2.29 (m, 2H), 2.02 (dd, J=16.32, 10.14 Hz, 2H), 1.83-1.96(m, 4H), 1.62-1.82 (m, 6H), 1.45-1.61 (m, 8H), 1.28-1.44 (m, 7H), 1.23(d, J=6.62 Hz, 4H), 1.07-1.19 (m, 7H), 1.04 (d, J=7.06 Hz, 6H). LCMS(ESI): m/z: [M+Na] calcd for C₅₃H₈₅N₃O₁₆Na: 1042.6; found 1042.6.

Example 50. Synthesis of Compound 2-CQ

Compound 2-CQ was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-(4-phenylpiperazin-1-yl)ethan-1-amine. ¹H NMR (400 MHz,Methanol-d4+Py-d5): δ ppm 6.97-7.06 (m, 1H), 6.87 (d, J=8.38 Hz, 3H),6.76 (t, J=7.06 Hz, 1H), 6.10-6.59 (m, 12H), 5.52 (d, J=5.29 Hz, 1H),4.76 (s, 1H), 4.65 (br. s., 1H), 4.52 (t, J=10.36 Hz, 1H), 4.23-4.42 (m,2H), 4.03-4.18 (m, 2H), 3.77-3.91 (m, 1H), 3.59-3.76 (m, 1H), 3.34-3.49(m, 4H), 3.18-3.26 (m, 1H), 2.96-3.14 (m, 5H), 2.77 (d, J=7.50 Hz, 1H),2.30-2.65 (m, 11H), 2.16-2.28 (m, 2H), 1.96-2.10 (m, 1H), 1.63-1.93 (m,6H), 1.36-1.61 (m, 6H), 1.34 (br. s., 3H), 1.23 (d, J=6.17 Hz, 3H), 1.12(d, J=6.17 Hz, 7H). LCMS (ESI): m/z: [M+H] calcd for C₅₉H₉₂N₅O₁₆:1126.65; found 1126.6.

Example 51. Synthesis of Compound 2-CR

Compound 2-CR was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with cyclopentylamine. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.03-6.60 (m, 13H), 5.48 (d,J=5.29 Hz, 2H), 5.35 (dd, J=14.33, 10.36 Hz, 2H), 4.68 (s, 1H), 4.59(br. s., 1H), 4.47 (t, J=9.92 Hz, 1H), 4.29 (t, J=9.48 Hz, 1H),4.15-4.24 (m, 1H), 3.91-4.09 (m, 3H), 3.80 (t, J=9.92 Hz, 1H), 3.69 (d,J=10.58 Hz, 1H), 3.55 (t, J=9.92 Hz, 1H), 3.33-3.40 (m, 2H), 3.20 (br.s., 1H), 2.74 (d, J=8.38 Hz, 1H), 2.27-2.54 (m, 4H), 2.13-2.25 (m, 2H),1.60-2.04 (m, 10H), 1.35-1.58 (m, 11H), 1.26-1.34 (m, 5H), 1.21 (d,J=6.62 Hz, 3H), 1.10 (d, J=6.17 Hz, 3H), 1.02 (d, J=7.06 Hz, 3H). LCMS(ESI): m/z: [M+H] calcd for C₅₂H₈₃N₃O₁₅Na: 1028.6; Found: 1028.6.

Example 52. Synthesis of Compound 2-CS

Compound 2-CS was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with4-tetrazolo-piperidine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm6.06-6.64 (m, 14H), 5.51 (br. s., 1H), 4.75 (s, 1H), 4.40-4.54 (m, 2H),4.28-4.39 (m, 3H), 4.24 (d, J=3.53 Hz, 1H), 3.96 (d, J=13.23 Hz, 1H),3.65-3.87 (m, 4H), 3.38-3.56 (m, 2H), 3.23 (d, J=9.70 Hz, 1H), 3.14 (br.s., 1H), 2.84 (t, J=12.13 Hz, 1H), 2.28-2.52 (m, 2H), 2.16-2.27 (m, 2H),1.78-2.08 (m, 8H), 1.62-1.77 (m, 3H), 1.29-1.61 (m, 6 H), 1.26 (d,J=6.17 Hz, 3H), 1.22 (d, J=6.17 Hz, 3H), 1.10 (d, J=6.62 Hz, 3H), 1.03(d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd for C₅₃H₈₃N₇O₁₆Na:1096.59; found 1096.6.

Synthesis of 4-tetrazolo-piperidine

Step 1: NH₄C₁ (3.28 g, 61.41 mmol, 3.00 equiv.) and NaN₃ (3.99 g, 61.41mmol, 3.00 equiv.) were added to a solution of compound 1 (5.00 g, 20.47mmol, 1.00 equiv.) in DMF (50.00 mL) and the resulting mixture wasstirred at 100° C. for 15 hrs. The reaction mixture was poured into H₂O(300 mL), and extracted with EtOAc (200 mL*3). Combined the organicphases were washed with brine (100 mL*5), dried over Na₂SO₄, filtered.Concentration under reduced pressure resulted in 4.6 g of 2 as lightyellow oil which was submitted to the next step with out furtherpurification. ¹H NMR (400 MHz, CDCl₃): δ ppm 7.34-7.28 (m, 5H), 5.15 (s,1 H), 4.25-4.21 (m, 2H), 3.32-3.30 (m, 1H), 3.29-3.26 (m, 2H), 2.13-2.04(m, 2H), 1.88-1.79 (m, 2H). LCMS (ESI): m/z: [M+Na] calcd forC₁₄H₁₇N₅O₂Na; 310.14; found 310.0.

Step 2: A mixture of compound 2 (2.60 g, 9.05 mmol, 1.00 equiv.) andPd/C (600.00 mg, 50% H₂O) in EtOH (120.00 mL) was stirred at 20° C.under H₂ for 24 hrs. The resulting mixture was filtered through celite,washed with MeOH:H₂O (5:1, about 200 mL), and concentrated under reducedpressure to give 1.0 g of compound 3 as white solid. ¹H NMR (400 MHz,D20): δ ppm 3.49-3.46 (m, 2H), 3.29-3.26 (m, 1H), 3.21-3.15 (m, 2H),2.27-2.24 (m, 2H), 2.05-1.96 (m, 2 H). LCMS (ESI): m/z: [M+H] calcd forC₆H₁₂N₅: 153.10; found 154.1.

Example 53. Synthesis of Compound 2-AW

Compound 2-AW was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted withN-(3-propylamino)morpholine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm8.67-8.85 (m, 1H), 6.10-6.61 (m, 12H), 5.55 (d, J=6.53 Hz, 1 H), 4.86(s, 1H), 4.64-4.77 (m, 1H), 4.55 (t, J=10.29 Hz, 1H), 4.27-4.45 (m, 2H),4.08-4.22 (m, 1H), 3.88 (t, J=9.54 Hz, 1H), 3.64-3.80 (m, 2H), 3.59 (t,J=4.27 Hz, 5H), 3.43-3.52 (m, 1H), 3.37 (br. s., 1H), 3.12-3.30 (m, 4H),2.59 (dd, J=14.31, 4.77 Hz, 1H), 2.35-2.54 (m, 2H), 2.18-2.33 (m, 9H),2.00-2.14 (m, 1H), 1.81-1.97 (m, 3H), 1.44-1.79 (m, 10H), 1.30-1.41 (m,4H), 1.22-1.30 (m, 3H), 1.14 (d, J=6.53 Hz, 3H), 1.07 (d, J=7.03 Hz,3H). LCMS (ESI): m/z: [M+H] calcd for C₅₄H₈₉N₄O₁₇: 1065.61; found1065.7.

Example 54. Synthesis of Compound 2-AT

Compound 2-AT was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with(1S,4S)-2-oxa-5-azabicyclo[2.2.1]heptane. ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 6.14-6.63 (m, 12H), 5.54 (d, J=5.02 Hz, 2H),4.76 (s, 1 H), 4.61-4.71 (m, 2H), 4.46-4.59 (m, 2H), 4.31-4.45 (m, 2H),4.18-4.28 (m, 1H), 4.16 (br. s., 1H), 3.96 (d, J=7.53 Hz, 1H), 3.86 (t,J=10.04 Hz, 1H), 3.70-3.82 (m, 2H), 3.45-3.53 (m, 1H), 3.26 (d, J=10.04Hz, 1H), 3.16 (s, 1H), 2.96 (d, J=8.03 Hz, 1H), 2.42-2.53 (m, 2H),2.35-2.41 (m, 1H), 2.21-2.31 (m, 2H), 1.98-2.09 (m, 1H), 1.64-1.97 (m,8H), 1.42-1.63 (m, 5H), 1.31-1.39 (m, 4H), 1.26 (d, J=6.53 Hz, 3H), 1.15(d, J=6.02 Hz, 3H), 1.07 (d, J=7.53 Hz, 3H). LCMS (ESI): m/z: [M+Na]calcd for C₅₂H₈₁N₃O₁₇Na: 1042.56; found 1042.6.

Example 55. Synthesis of Compound 2-CL

Compound 2-CL was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with allylpiperidine-4-carboxylate.

Deprotection Step. To a solution of allyl carboxylate of 2-CL (500.00mg, 458.59 umol, 1.00 equiv.) in DMF (13.00 mL) was added2-sulfanylbenzoic acid (141.42 mg, 917.18 umol, 2.00 equiv.) was addedPd(PPh₃)₄ (264.96 mg, 229.29 umol, 0.50 equiv.) under N₂ protected. Themixture was stirred at 25° C. for 1 hr, and filtered. The resultingmixture was purified by prep-HPLC (FA) chromatography to give 50.00 mgof 2-CL as a light yellow solid. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δppm 6.03-6.56 (m, 13H), 5.47 (d, J=7.06 Hz, 1H), 5.35 (dd, J=14.11,10.58 Hz, 1H), 4.72 (s, 1H), 4.39-4.57 (m, 1H), 4.25-4.37 (m, 1H),4.10-4.25 (m, 2H), 3.75-3.89 (m, 1H), 3.57-3.73 (m, 2H), 3.39-3.48 (m,1H), 3.32-3.38 (m, 1H), 3.17-3.25 (m, 1H), 2.73-2.89 (m, 2H), 2.29-2.49(m, 3H), 2.14-2.25 (m, 1H), 1.79-1.92 (m, 5H), 1.59-1.76 (m, 6H),1.29-1.58 (m, 6H), 1.26 (d, J=5.73 Hz, 3H), 1.21 (d, J=6.17 Hz, 3H),1.10 (d, J=6.62 Hz, 3H), 1.03 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z:[M+Na] calcd for C₅₃H₈₅N₃O₁₈Na: 1072.59; found 1072.6.

Example 56. Synthesis of Compound 2-CT

Compound 2-CT was synthesized in the manner similar to Compound 2-I(Example 3) and 2-CL (Example 55), except piperazine was substitutedwith allyl-4-(aminomethyl)benzoate. Two peaks were isolated upondeprotection and prep-HPLC purification. Peak one: ¹H NMR (400 MHz,Methanol-d4+Pyr-d5): δ ppm 8.03 (d, J=7.50 Hz, 2H), 7.32 (d, J=7.06 Hz,2H), 6.06-6.52 (m, 11H), 4.75 (s, 1H), 4.60-4.72 (m, 1H), 4.48 (d,J=16.32 Hz, 1H), 4.26-4.41 (m, 2H), 4.08-4.25 (m, 1H), 3.86 (br. s.,1H), 3.62-3.81 (m, 3H), 3.47 (br. s., 1H), 3.16 (br. s., 1H), 2.33-2.52(m, 3H), 2.19-2.27 (m, 2H), 2.03 (br. s., 1H), 1.89 (d, J=14.11 Hz, 2H),1.64-1.78 (m, 2H), 1.33-1.62 (m, 6H), 1.30 (d, J=5.73 Hz, 3H), 1.23 (d,J=6.17 Hz, 3H), 1.11 (d, J=6.17 Hz, 3H), 1.04 (d, J=6.62 Hz, 3H). LCMS(ESI): m/z: [M+H] calcd for C₅₅H₈₂N₃O₁₈: 1072.55; found 1072.50. Peak 2:¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm 8.10 (d, J=6.62 Hz, 3H), 7.36(d, J=7.50 Hz, 3H), 5.82-6.78 (m, 9H), 4.93 (br. s., 1H), 4.60-4.84 (m,2H), 4.45 (br. s., 3H), 4.22 (br. s., 2H), 3.81-4.11 (m, 4H), 3.70 (br.s., 2H), 2.42-2.60 (m, 3H), 2.25 (br. s., 1H), 1.40-2.14 (m, 14H),1.19-1.36 (m, 7H), 1.10 (br. s., 4 H), 0.92-1.06 (m, 3H). LCMS (ESI):m/z: found 1054.40.

Example 57. Synthesis of Compound 2-AR

Compound 2-AR was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with8-oxa-3-azabicyclo[3.2.1]octane. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δppm 6.05-6.51 (m, 8H), 5.47 (br. s., 1H), 5.34-5.43 (m, 3H), 4.79 (s,1H), 4.61 (br. s., 1H), 4.46-4.54 (m, 1H), 4.36 (d, J=9.26 Hz, 1H),4.10-4.27 (m, 3H), 3.76-3.87 (m, 1H), 3.60-3.75 (m, 3H), 3.37-3.45 (m,1H), 3.33 (d, J=3.97 Hz, 1H), 3.24 (br. s., 2H), 2.99 (d, J=12.35 Hz,2H), 2.29-2.46 (m, 1H), 2.14-2.28 (m, 1H), 1.79-1.96 (m, 2H), 1.60-1.78(m, 5H), 1.49-1.59 (m, 3H), 1.37-1.49 (m, 2H), 1.16-1.32 (m, 6H),1.00-1.13 (m, 6H). LCMS (ESI): m/z: [M+Na] calcd for C₅₃H₈₃N₃O₁₇Na:1056.57; found 1056.6.

Example 58. Synthesis of Compound 2-CU

Compound 2-CU was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with methan-d3-amine. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.20-6.70 (m, 13H), 5.64 (d,J=6.17 Hz, 2H), 4.95 (s, 1H), 4.79 (d, J=5.73 Hz, 1H) 4.64 (t, J=10.36Hz, 1H,) 4.34-4.55 (m, 3H), 4.14-4.32 (m, 1H), 3.97 (br. s., 1H),3.71-3.91 (m, 3H), 3.58 (dd, J=9.26, 6.17 Hz, 2H), 3.38 (d, J=9.26 Hz,1H), 3.28 (d, J=7.50 Hz, 1H), 2.92-3.08 (m, 1H), 2.79 (d, J=6.62 Hz,1H), 2.45-2.69 (m, 3H), 2.24-2.42 (m, 2H) 2.09-2.22 (m, 1H), 1.90-2.06(m, 3H), 1.75-1.88 (m, 3H), 1.52-1.73 (m, 6H), 1.42-1.51 (m, 4H), 1.37(d, J=6.17 Hz, 3H), 1.25 (d, J=5.73 Hz, 3H), 1.18 (d, J=7.06 Hz, 3 H).LCMS (ESI): m/z: [M+Na] calcd for C₄₈H₇₄D₃N₃O₁₆Na: 977.55; found 977.5.

Example 59. Synthesis of Compound 2-CJ

Compound 2-CJ was synthesized in the manner similar to Compound 2-I(Example 3) and 2-CL (Example 55), except piperazine was substitutedwith allyl (3-aminopropyl)carbamate. LCMS (ESI): m/z: [M+H] found 995.5.

Example 60. Synthesis of Compound 2-CK

Compound 2-CK was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with6-oxa-2-azaspiro[3.4]octane. LCMS (ESI): m/z: [M+Na] found 1056.3.

Example 61. Synthesis of Compound 2-Q

Compound 2-Q was synthesized in the manner similar to Compound 2-I(Example 3), except piperazine was substituted with2-(1H-1,2,4-triazol-1-yl)ethan-1-amine. LC-MS: Calculated(C₅₁H₅₀N₆O₁₆+H)⁺: 1033.56. Observed: 1033.50.

Example 62. Sample Characterization Data of Ureas 2

TABLE 2 Sample Characterization Data of Ureas 2 Isolated LC Yieldretention Compound from Observed time LC LC ID 1-2 Mass (min) ConditionsPurity 2-B  7% 988.6 (M + Na) 7.39 A 98% 2-C  3% 1028.6 (M + Na) 7.88 A97% 2-E  6% 1030.6 (M + Na) 7.46 A 99% 2-J 10% 1043.6 (M + Na) 6.81 A97% 2-D 24% 1000.5 (M + Na) 7.4 A 97% 2-F 13% 1009.6 (M + H) 6.87 A 96%2-I 23% 1029.6 (M + Na) 6.8 A 98% 2-T 10% 1065.6 (M + Na) 7.07 A 99% 2-U18% 1051.6 (M + Na) 6.96 A 99% 2-Y 23% 1018.5 (M + Na) 7.37 A 99% 2-AG 6% 1014.5 (M + Na) 7.97 A 99% 2-AB  8% 1071.6 (M + Na) 7.55 A 99% 2-AF 9% 1051.7 (M + H) 7.41 A 99% 2-AI 19% 1014.5 (M + Na) 7.55 A 99% 2-BC38% 1033.4 (M + Na) 4.55 C 98%

Conditions for LC analysis: Conditions A: Agilent 1260, 6120 MS, Column:Phenomenex Luna μm C18(2) 100 A 50×2.0 mm, 0.8 mL/min, columntemperature: 40° C., mobile Phase: A: 4 L H₂O (with 1.5 mL TFA) B: 4 LAcetonitrile (with 0.75 mL TFA), Gradient (min, % B): 0, 10; 0.4, 10;3.40, 100; 3.85, 100; 3.86, 10. LC purity calculated from the peak arearatio monitoring at 383 nM. Conditions B: Agilent LCMS, Zorbax EclipseC18 1.8 μM, 2.1×50 mm, 0.4 mL/min, linear gradient from 95:5 to 5:95H₂O, acetonitrile over 8 minutes with each eluent containing 0.1% formicacid. Conditions C: Shimadzu LC-MS system (Shimadzu Co., Japan),Phenomenex Onyx Monolithic C18 column (4.6×50 mm), p/n CHO-7644(Phenomenex Co.); samples dissolved in DMSO were eluted using a lineargradient of 0.1% HCOOH in 100% water (mobile phase A) to 0.1% HCOOHacetonitrile in 100% acetonitrile (mobile phase B).

Example 63. C16-Carbamates 3

Preparation of 3-2: To a 40 mL vial was added 3-1 (602.6 mg, 275.3 μmol,1 eq.), prepared as described in Driver et al., J Chem Soc Perkin Trans3155-7 (1992), and benzene (13.7 mL). Triethylamine (115 μL, 0.822 mmol,3 eq.) was added followed by DPPA (71 μL, 33.0 mmol, 1.2 eq.). Thereaction was then placed in a preheated heating block at 80° C. andallowed to stir for 3.5 hours. The reaction was then transferred to a125 mL separatory funnel with water (25 mL) and diethyl ether (50 mL).The layers were separated and the organic layer was washed with brine(25 mL), dried over Na₂SO₄, filtered and concentrated in vacuo. Theresulting red/orange oil was then purified by SiO₂ chromatography (100:0to 0:100 Hexane:Et₂O) yielding 3-2 as an orange solid (168.7 mg, 0.077mmol, 28% yield). TLC (hexanes: Et₂O 7:3) R_(f)=0.64, visualized by CAMHRMS (ESI): Calculated for C₁₁₇H₂₁₀N₂O₁₈Si₉ (M+Na⁺): 2206.3400, Found:2206.3413.

Preparation of 3-3-A: To a 1.5 mL vial was added 3-2 (as a stocksolution (100 μL of 150 mg in 1.5 mL benzene) 10 mg, 4.57 μmol, 1 eq.),and titanium isopropoxide (as a stock solution (50 μL of 25 μL in 4.6 mLbenzene) 0.27 μL, 0.914 μmol, 0.2 eq.) and THF (80 μL). The reaction wasthen allowed to stir at room temperature for 1 h. The reaction was thendiluted with water (1.5 mL) and diethyl ether (1.5 mL). The layers wereseparated and the organic layer was dried over Na₂SO₄, filtered andconcentrated in vacuo. The resulting red/orange oil was then purified bySiO₂ chromatography (100:0 to 80:20 hexane:Et₂O) yielding 3-3-A as anorange solid. TLC (hexanes: Et₂O 7:3), R_(f)=0.51, stained by CAM. LRMS(ESI) 2266.6 (M+Na).

¹H NMR (500 MHz, Acetone-d6): δ 7.88 (d, J=7.5 Hz, 2H), 7.70 (d, J=7.5Hz, 2H), 7.43 (t, J=7.3 Hz, 2H), 7.36-7.32 (m, 2H), 6.59-6.08 (m, 12H),6.03 (dd, J=15.5, 6.1 Hz, 1H), 5.51 (dd, J=14.9, 9.5 Hz, 1H), 5.35 (d,J=9.9 Hz, 1H), 4.87 (p, J=6.3 Hz, 1H), 4.77-4.73 (m, 1H), 4.71-4.67 (m,1H), 4.65 (s, 1H), 4.48 (dd, J=10.5, 6.5 Hz, 1H), 4.37 (dd, J=10.4, 6.5Hz, 1H), 4.25 (app t, J=6.3 Hz, 2H), 4.18-4.09 (m, 1H 4.07-3.97 (m, 2H),3.87-3.84 (m, 1H), 3.76 (app dd, J=11.8, 6.9 Hz, 1H), 3.70 (d, J=8.9 Hz,1H), 3.74-3.66 (m, 2H), 3.47-3.34 (m, 2H), 3.35-3.28 (m, 1H), 3.15 (s,3H), 2.58 (d, J=6.6 Hz, 1H), 2.47-2.40 (m, 2H), 2.26 (app dd, J=15.6,7.4 Hz, 2H), 2.20-2.15 (m, 1H), 1.94-1.85 (m, 4H), 1.84-1.80 (d, J=13.1Hz, 3H), 1.79-1.68 (m, 4H), 1.68-1.61 (d, J=9.3 Hz, 2H), 1.54-1.56 (s,1H), 1.26 (app dd, J=6.2, 3.2 Hz, 6H), 1.22 (d, J=6.2 Hz, 3H), 1.18 (d,J=6.0 Hz, 3H), 1.14-0.83 (m, 87H), 0.81-0.53 (m, 54H).

Preparation of 3-A: Treatment of 3-3-A with HF, pyridine, evaporation ofthe solution and treatment of the residue with piperidine in DMFprovides the carbamate 3-A after purification by HPLC with 0.1% to 0.3%formic acid modified H₂O/CH₃CN.

Specific Compounds 3

where HOR′ is depicted in Table 3:

TABLE 3

A

B

C

D

E

F

G

H

I

J

K

L

M

N

O

Example 64. C16 Amides 4

Amides 4 are prepared according to the routes described in Scheme 4.Specifically, treatment of the isocyanate 3-2 with organometallicreagents (i.e., organozinc reagents, organomagnesium reagents,organolithium reagents) provides the desired amides intermediates 4-1according, for example, to the methods described in Carlin and Smith, JAm Chem Soc 69: 2007 (1947) or Szczesniak et al., J Org Chem 79(23):11700-13 (2014). Deprotection of the amphotericin derivative isaccomplished generally according to the methods of Driver et al., J ChemSoc Perkin Trans 3155-7 (1992) allows the isolation of the desiredamides 4, as described in Scheme 4.

Specific Compounds 4

where R′ and the site of connectivity are depicted in Table 4:

TABLE 4

A

B

C

D

E

F

G

H

I

J

Example 65. C3′ Amides and Carbamates 5

Compounds 5 are accessed from 2-BF (Example 4) according to theprocedure established by Wright et al., J Antibiotics 35: 911-4 (1982).Specifically, for the synthesis of 5-C, the bis-Fmoc-protectedN-hydroxysuccinimide ester of D-lysine (3 equiv., prepared as describedin Russ, J Bioorg Chem 33: 139 (2007)) and Et₃N (1 equiv.) are added toa solution of 2-BF (1 equiv.) in dry DMF. The reaction mixture is keptat 37° C. for 1 h, and then H₂O is added. The mixture is extracted withbutanol. Organic fractions are combined concentrated. The addition ofdiethyl ether provides a yellow precipitate, which is filtered off,washed with diethyl ether and purified to produce 5-C. In examples wherethe Fmoc protecting group is not cleaved from the substrate during thecoupling reaction, it can be efficiently removed using1,5-diazabicyclo[4.3.0]non-5-ene (DBN) or piperidine in DMF, followed byreprecipitation by addition of the DMF mixture to a large volume ofdiethyl ether.

Alternatively, Compounds 5 can be prepapered by the following procedure:

Specific Compounds 5

where R₃ defined according to the structure above are depicted in Table5, and XR₁ defined according to the structure above are depicted inTable 6:

TABLE 5

A

B

C

D

E

F

G

H

I

J

TABLE 6

Q

R

S

T

U

Example 66. Synthesis of Compound 5-DT

Step. 1: The mixture of compound(R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(pyridin-3-yl)propanoicacid (301.00 mg, 774.94 umol, 1.65 equiv.) and HOBt (126.92 mg, 939.32umol, 2.00 equiv.) in DMF (0.6 mL) was prepared and cooled to 0° C., DCC(145.36 mg, 704.49 umol, 142.51 uL, 1.50 equiv.) was added. The reactionmixture was stirred at 0° C. for 1 hr, the residue of DCU was filtered,the obtained eluate was added to the solution of 2-AG (Example 25)(466.00 mg, 469.66 umol, 1.00 equiv.) in DMF (5.00 mL), then DIPEA(242.80 mg, 1.88 mmol, 328.11 uL, 4.00 equiv.) was added to the reactionmixture dropwise. The reaction was stirred at r.t. for 28 hrs. Themixture was poured into MTBE (150 mL), and then filtered to give 426 mgof intermediate 5-1 as yellow solid which was used to next step.

Step 2: The mixture of intermediate 5-1 (247.00 mg, 181.27 μmol, 1.00equiv.) in DMSO (3.00 mL) was added piperidine (154.35 mg, 1.81 μmol,0.01 equiv.) at r.t. The mixture was stirred at r.t. for 0.2 hr. Themixture was filtered and purified by prep-HPLC (FA) to give 31.40 mg ofCompound 5-DT as yellow solid. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δppm 8.58-8.68 (m, 2H), 8.24-8.34 (m, 2H), 7.67-7.72 (m, 1H), 7.24-7.27(m, 1H), 6.13-6.45 (m, 13H), 5.09-5.24 (m, 2H), 4.71 (s, 1H), 4.27-4.70(m, 2H), 4.00-4.27 (m, 3H), 3.79-4.00 (m, 3H), 3.76-3.79 (m, 2H),3.29-3.76 (m, 3H), 3.19-3.29 (m, 5H), 2.18-2.12 (m, 9H), 1.81-1.83 (m,5H), 1.52-1.55 (m, 9H), 1.3 (d, J=8 Hz, 3H), 1.20 (d, J=8 Hz, 3 H), 1.10(d, J=8 Hz, 3H), 1.02 (d, J=8 Hz, 3H). LCMS (ESI): m/z: [M+H] calcd forC₅₉H₉₀N₅O₁₇: 1140.63; found 1140.60.

Example 67. Synthesis of Compound 5-DR

Compound 5-DR was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-BG. ¹H NMR(400 MHz, Methanol-d4+Pyr-d5): δ ppm 8.53 (s, 1H), 8.34 (d, J=3.53 Hz,1H), 7.76 (d, J=7.50 Hz, 1H), 7.16-7.19 (m, 1H), 6.04-6.58 (m, 13H),5.30-5.54 (m, 3H), 4.79 (s, 1H), 4.46-4.67 (m, 1H), 4.24-4.39 (m, 2H),4.04-4.22 (m, 3H), 3.41-3.87 (m, 6H), 3.06-3.28 (m, 5H), 2.55 (d,J=11.03 Hz, 1H), 2.30-2.50 (m, 2H), 2.15-2.29 (m, 2H), 1.36-2.11 (m,11H), 1.33 (d, J=6.17 Hz, 3 H), 1.23 (d, J=6.17 Hz, 3H), 1.12 (d, J=6.62Hz, 3H), 1.04 (d, J=7.50 Hz, 3H). LCMS (ESI): m/z: [M+H] calcd forC₅₇H₈₉N₆O₁₇: 1129.62; found: 1129.60.

Example 68. Synthesis of Compound 5-DS

Compound 5-DS was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-AF(Example 15). ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm 8.34 (d, J=4.85Hz, 1H), 7.74 (s, 4H), 7.72 (s, 4H), 7.69 (br. s., 1H), 7.46 (s, 1H),7.20-7.23 (m, 2H), 6.37-6.55 (m, 2H), 6.24-6.37 (m, 7H), 6.01-6.24 (m,4H), 5.48 (d, J=6.17 Hz, 1H), 5.35 (dd, J=14.33, 9.92 Hz, 1H), 4.75 (s,1H), 4.61 (br. s., 1 H), 4.48 (t, J=9.92 Hz, 1H), 4.14-4.35 (m, 1H),3.97-4.10 (m, 2H), 3.92 (s, 4H), 3.74-3.88 (m, 1H), 3.48-3.72 (m, 8H),3.32-3.47 (m, 2H), 3.05-3.27 (m, 4H), 2.80-3.00 (m, 1H), 2.37-2.47 (m,31H), 2.13-2.26 (m, 2H), 1.84 (d, J=4.85 Hz, 2H), 1.65-1.80 (m, 2H),1.54-1.65 (m, 4H), 1.46-1.53 (m, 17H), 1.29-1.38 (m, 12H), 1.21 (d,J=6.17 Hz, 3H), 1.07-1.12 (m, 4H), 1.02 (d, J=7.06 Hz, 3H). LCMS (ESI):m/z: [M+H] calcd for C₆₁H₉₅N₆O₁₈: 1200.66; found: 1200.60.

Example 69. Synthesis of Compound 5-QA

Compound 5-QA was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-BF(Example 4); and(R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(pyridin-3-yl)propanoicacid with NN-dimethylglycine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δppm 6.21-6.79 (m, 13H), 5.73 (d, J=5.73 Hz, 1H), 5.03 (s, 1H), 4.86 (br.s., 1H), 4.73 (t, J=10.14 Hz, 1H), 4.48-4.62 (m, 2 H), 4.24-4.43 (m,3H), 4.04 (t, J=9.70 Hz, 1H,) 3.82-3.99 (m, 3H), 3.69 (dd, J=8.82, 6.17Hz, 1H), 3.53 (d, J=10.14 Hz, 1H), 3.44-3.49 (m, 4H) 3.41 (d, J=9.26 Hz,1H), 2.98-3.15 (m, 2H), 2.76-2.97 (m, 4H), 2.50-2.70 (m, 2H), 2.36-2.48(m, 2H), 2.24 (s, 7H), 2.02-2.15 (m, 2H), 1.82-2.00 (m, 4H), 1.63-1.80(m, 5H) 1.44-1.62 (m, 5H), 1.41 (d, J=6.62 Hz, 3H), 1.28 (d, J=6.62 Hz,3H), 1.22 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd forC₅₂H₈₄N₄O₁₇Na: 1059.58; found 1059.5.

Example 70. Synthesis of Compound 5-QB

Compound 5-QB was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-BF(Example 4) and(R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(pyridin-3-yl)propanoicacid with fmoc-glycine. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm6.19-6.78 (m, 13H), 5.72 (d, J=5.73 Hz, 1H), 5.01 (s, 1H), 4.83 (br. s.,1H), 4.71 (t, J=9.92 Hz, 1H), 4.47-4.62 (m, 2H), 4.24-4.44 (m, 3 H),4.03 (t, J=9.48 Hz, 1H), 3.81-3.97 (m, 3H), 3.75 (s, 1H), 3.68 (dd,J=8.60, 6.39 Hz, 1H), 3.53 (br. s., 1H), 3.40 (d, J=9.26 Hz, 1H),2.76-3.02 (m, 4H), 2.50-2.69 (m, 2H), 2.33-2.47 (m, 2H), 2.25 (d,J=10.58 Hz, 1H), 2.05 (d, J=6.62 Hz, 2H), 1.80-1.99 (m, 4H), 1.62-1.79(m, 5H), 1.51 (d, J=5.73 Hz, 5H), 1.40 (d, J=6.17 Hz, 3H), 1.28 (d,J=6.17 Hz, 3H), 1.21 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcdfor C₅₀H₅₀N₄O₁₇Na: 1031.55; found 1031.6.

Example 71. Synthesis of Compound 5-QC

Compound 5-QC was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-BF(Example 4) and(R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(pyridin-3-yl)propanoicacid with(R)-2,5-bis((((9H-fluoren-9-yl)methoxy)carbonyl)amino)pentanoic acid. ¹HNMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.12-6.69 (m, 13H), 5.61 (br.s., 1H), 5.44-5.52 (m, 2H), 4.87 (s, 1H), 4.71 (br. s., 1H), 4.60 (t,J=10.36 Hz, 1H), 4.30-4.48 (m, 2H), 4.09-4.25 (m, 2H), 3.93 (t, J=9.48Hz, 1H), 3.64-3.85 (m, 3H), 3.50-3.61 (m, 1H), 3.46 (br. s., 1H),3.31-3.37 (m, 1H), 3.13 (br. s., 2H), 2.84 (s, 3H), 2.42-2.68 (m, 2H),2.26-2.38 (m, 2H), 1.72-2.15 (m, 10H), 1.50-1.70 (m, 5H), 1.44 (d,J=5.73 Hz, 4H), 1.34 (d, J=6.17 Hz, 3H), 1.23 (d, J=6.17 Hz, 3H), 1.15(d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+H] calcd for C₅₃H₈₈N₅O₁₇:1066.61; found 1066.6.

Example 72. Synthesis of Compound 5-QD

Compound 5-QD was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-BF(Example 4). ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm 8.63-8.65 (m,1H), 8.43-8.53 (m, 1H), 7.80 (d, J=7.94 Hz, 1 H), 7.21-7.28 (m, 1H),6.21-6.73 (m, 13H), 5.69 (d, J=5.29 Hz, 1H), 4.96 (s, 1H), 4.80 (br. s.,1H), 4.68 (t, J=10.36 Hz, 1H), 4.39-4.57 (m, 2H), 4.19-4.36 (m, 3H),3.95-4.07 (m, 2 H), 3.86-3.92 (m, 2H), 3.75-3.85 (m, 1H), 3.64 (dd,J=8.82, 6.17 Hz, 1H), 3.50 (br. s., 1H), 3.38 (d, J=9.70 Hz, 1H), 3.30(dd, J=14.11, 4.85 Hz, 1H), 3.06 (dd, J=13.67, 7.50 Hz, 1H), 2.91 (s,3H), 2.81 (d, J=11.47 Hz, 1H), 2.47-2.67 (m, 2H), 2.33-2.45 (m, 2H),2.14-2.28 (m, 1H), 1.98-2.11 (m, 2H), 1.78-1.97 (m, 3H), 1.59-1.77 (m,5H), 1.43-1.58 (m, 5H), 1.38 (d, J=6.17 Hz, 3H), 1.26 (d, J=6.62 Hz,3H), 1.19 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd forC₅₆H₈₅N₅O₁₇Na: 1122.59; found 1122.5.

Example 73. Synthesis of Compound 5-UA

Compound 5-UA was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-J (Example8), and(R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(pyridin-3-yl)propanoicacid with NN-dimethylglycine. ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm6.28-6.62 (m, 14H), 5.47-5.45 (m, 1H), 5.36-5.41 (m, 1H), 4.87 (s, 1H),4.70-4.75 (m, 1H), 4.58-4.64 (t, 1H), 4.41-4.50 (m, 2H), 4.25-4.35 (m,1H), 4.20-4.25 (m, 1H), 4.14 (d, J=2.8 Hz, 1H), 3.80-3.95 (m, 3H),3.66-3.86 (m, 4 H), 3.30-3.46 (m, 1H), 3.36-3.41 (m, 1H), 3.20 (s, 2H),2.36-2.64 (m, 17H), 1.52-2.12 (m, 13 H), 1.33-1.35 (m, 4H), 1.34 (d,J=6.4 Hz, 3H), 1.23 (d, J=6.4 Hz, 3H), 1.25 (d, 3H) 1.15 (d, 3 H). LCMS(ESI): m/z: [M+Na] calcd for C₅₆H₉₁N₅O₁₇Na: 1128.6; found 1128.5.

Example 74. Synthesis of Compound 5-UB

Compound 5-UB was synthesized in the manner similar to Compound 5-DT(Example 66), except 2-AG (Example 25) was substituted with 2-J (Example8), and(R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(pyridin-3-yl)propanoicacid with fmoc-glycine. ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ 6.26-6.59(m, 14H), 5.36-5.44 (m, 2H), 4.85 (s, 1H), 4.68-4.72 (m, 1H), 4.55-4.63(t, 1H), 4.38-4.43 (m, 2H), 4.20-4.30 (m, 2H), 4.13-4.15 (m, 1 H),3.80-3.93 (m, 4H), 3.52-3.68 (m, 4H), 3.36-3.46 (m, 4H), 2.45-2.49 (m,5H), 2.27-2.34 (m, 6H), 1.52-2.12 (m, 13H), 1.42-1.45 (m, 4H), 1.33 (d,J=6.0 Hz, 3H), 1.22 (d, J=6.0 Hz, 3 H), 1.42 (d, J=7.2 Hz, 3H). LCMS(ESI): m/z: [M+Na] calcd for C₅₄H₈₇N₅O₁₇Na: 1100.61; found 1100.5.

Example 75. C3′ Derivatives 6

C3′ alkyl derivatives 6 are prepared according to the procedure definedby Paquet et al., Chem Eur J 14: 2465-81 (2008). Specifically, treatmentof 2-BF (Example 4) with 1H-pyrazole-1-carboxamidine monohydrochloride(1 equiv.) and diisopropylethylamine (3 equiv.) in DMF at roomtemperature provides the guanidine compound 6-E. Analog 6-B issynthesized by treatment of 2-BF withN-(9-fluorenylmethoxycarbonyl)-3-aminopropanal (4 equiv.) and NaBH₃CN (4equiv.) in DMF with catalytic HCl. Filtration and precipitation byaddition to diethyl ether provides a yellow precipitate that can bepurified by normal or reverse phase chromatography. Dissolution of thisintermediate in DMF and treatment with piperidine (8 equiv.) at roomtemperature, followed by precipitation by addition to diethyl ether,provides 6-B.

Specific Compounds 6

partial structures of which, defined according to the structure above,are depicted in Table 7 and Table 8:

TABLE 7 6 N R4, R5

A

B

C

D

E

TABLE 8 6 XR1

Q

R

S

T

U

Example 76. Synthesis of Compound 6-QB

Step 1: To a solution of (9H-fluoren-9-yl)methyl (3-oxopropyl)carbamate(775.45 mg, 2.63 mmol, 5.00 equiv.) and 2-BF (Example 4) (500.00 mg,525.14 umol, 1.00 equiv.) in DMF (15.00 mL) was added NaBH(OAc)₃ (1.11g, 5.25 mmol, 10.00 equiv.) at r.t. for 1.5 hours. The mixture waspoured into MTBE (200 mL) and filtered to give the solution of 3 g ofcrude (Fmoc)₂-6-QB which was used to next step directly.

Step 2: To the solution of compound (Fmoc)₂-6-QB (3 g, 1.99 mmol, 1.00equiv.) in DMSO (about 20 mL) was added Et₃N (2.01 g, 19.90 mmol, 10.00equiv.) and stirred at r.t. for 14 hrs. The reaction was poured intoMTBE (200 mL) and precipitate was filtered to give yellow solid that waspurified by prep-HPLC (FA) chromatography to yield 24.0 mg of 6-QB asyellow solid. ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.03-6.57 (m,12H), 4.68 (s, 1H), 4.59 (br. s., 1H), 4.49 (t, J=9.70 Hz, 1H),4.22-4.39 (m, 3H), 4.12 (d, J=9.70 Hz, 1H), 3.78-3.93 (m, 2H), 3.71 (d,J=10.58 Hz, 1H), 3.52 (t, J=6.39 Hz, 2H), 3.03-3.19 (m, 6H), 2.88-3.00(m, 2H), 2.83 (d, J=11.47 Hz, 2H), 2.68 (s, 3H), 2.29-2.54 (m, 4H), 2.21(d, J=16.32 Hz, 2H), 2.02 (d, J=5.73 Hz, 1H), 1.61-1.95 (m, 10H),1.38-1.62 (m, 7H), 1.32 (d, J=5.73 Hz, 4H), 1.22 (d, J=6.17 Hz, 3H),1.10 (d, J=6.17 Hz, 3H), 1.03 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+H]calcd for C₅₅H₉₂N₅O₁₆: 1066.5; found 1066.5.

Example 77. Synthesis of Compound 6-TB

Compound 6-TB was synthesized in the manner similar to Compound 6-QB(Example 76), except 2-BF (Example 4) was substituted with 2-AG (Example25). ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm 6.09-6.57 (m, 13H), 5.52(d, J=5.29 Hz, 1H), 4.45-4.74 (m, 4 H), 4.20-4.42 (m, 4H), 4.13 (d,J=3.53 Hz, 2H), 3.79-3.96 (m, 3H), 3.74 (d, J=10.58 Hz, 1 H), 3.56 (br.s., 1H), 3.39 (dd, J=8.38, 6.17 Hz, 1H), 3.24 (d, J=9.26 Hz, 2H),3.03-3.20 (m, 6 H), 2.90-3.02 (m, 2H), 2.86 (d, J=8.82 Hz, 1H),2.28-2.54 (m, 4H), 1.62-2.28 (m, 20H), 1.26-1.62 (m, 14H), 1.23 (d,J=6.17 Hz, 3H), 1.12 (d, J=6.62 Hz, 3H), 1.04 (d, J=7.06 Hz, 3 H). LCMS(ESI): m/z: [M+H] calcd for C₅₇H₉₆ N₅O₁₆: 1106.3; found 1106.7.

Example 78. Synthesis of Compound 6-UB

Compound 6-UB was synthesized in the manner similar to Compound 6-QB(Example 76), except 2-BF (Example 4) was substituted with 2-CR (Example51). ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.08-6.58 (m, 14H),5.52 (d, J=5.27 Hz, 1H), 4.72 (s, 1H), 4.63 (br. s., 1H), 4.52 (t,J=10.48 Hz, 1H), 4.24-4.43 (m, 4H), 4.01-4.16 (m, 3H), 3.80-3.97 (m,3H), 3.75 (d, J=10.54 Hz, 1H), 3.38-3.47 (m, 2H), 3.27 (d, J=8.16 Hz,1H), 3.08-3.21 (m, 5 H), 2.99 (dd, J=13.18, 6.27 Hz, 3H), 2.87 (d,J=10.79 Hz, 1H), 2.34-2.52 (m, 4H), 2.20-2.29 (m, 2H), 1.29-2.10 (m,35H), 1.25 (d, J=6.40 Hz, 3H), 1.14 (d, J=6.27 Hz, 3H), 1.07 (d, J=7.15Hz, 3H). LCMS (ESI): m/z: [M+H] calcd for C₅₈H₉₈ N₅O₁₆: 1020.4; found1020.7.

Example 79. Synthesis of Compound 6-SB

Compound 6-SB was synthesized in the manner similar to Compound 6-QB(Example 76), except 2-BF (Example 4) was substituted with 2-AF (Example15). ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm 8.69 (s, 2H), 6.01-6.61(m, 10H), 5.44-5.57 (m, 1H), 5.39 (br. s., 3H), 4.71 (s, 1H), 4.51 (br.s., 2H), 4.18-4.41 (m, 3H), 3.84 (dd, J=14.77, 9.48 Hz, 3H), 3.58 (br.s., 5H), 3.35 (dd, J=14.77, 7.72 Hz, 3H), 3.24 (d, J=9.26 Hz, 2H),3.06-3.20 (m, 5 H), 2.82-3.03 (m, 3H), 2.28-2.52 (m, 10H), 1.80-2.07 (m,8H), 1.39-1.76 (m, 9H), 1.33 (d, J=6.17 Hz, 3H), 1.22 (d, J=6.17 Hz,3H), 1.11 (d, J=6.17 Hz, 3H), 1.04 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z:[M+H] calcd for C₅₉H₁₀₁N₆O₁₆: 1165.71; found 1165.70.

Example 80. Synthesis of Compound 6-QE

To a solution of compound 2-BF (Example 4) (380.00 mg, 399.11 umol, 1.00equiv.) in DMF (4.00 mL) was added 1H-pyrazole-1-carboximidamide (109.87mg, 997.78 umol, 2.50 equiv.) followed by DIPEA (412.65 mg, 3.19 mmol,8.00 equiv.). The resulting mixture was stirred at RT for 40 hr.,filtered to give the filtrate which was purified by prep-HPLC (FA)chromatography to yield 36.00 mg of AmBMU-A3 as yellow solid. ¹H NMR(400 MHz, Methanol-d4+Py-d5): δ ppm 9.06 (s, 1H), 6.20-6.69 (m, 12H),5.66 (d, J=5.73 Hz, 1H), 4.93 (s, 1H), 4.78 (br. s., 1H), 4.67 (t,J=9.92 Hz, 1H), 4.42-4.55 (m, 2H), 4.22-4.37 (m, 2H), 3.93-4.05 (m, 2H),3.89 (d, J=11.03 Hz, 1H), 3.71-3.82 (m, 1H), 3.54-3.65 (m, 1H), 3.49(br. s., 1H), 3.36-3.42 (m, 1H), 3.16 (s, 1H), 2.76-2.90 (m, 3H),2.47-2.69 (m, 3H), 2.33-2.44 (m, 2H), 2.14-2.23 (m, 1H), 1.97-2.12 (m,2H), 1.81-1.93 (m, 2H), 1.48-1.77 (m, 6 H), 1.45 (d, J=5.73 Hz, 3H),1.38 (d, J=6.62 Hz, 3H), 1.26 (d, J=6.17 Hz, 3H), 1.19 (d, J=7.06 Hz,3H). LCMS (ESI): m/z: [M+H] calcd for C₄₉H₇₉N₅O₁₆: 994.15; found 994.5.

Example 81. C3′-azide 7

Compound 7 is prepared starting from intermediate 3-2. Treatment of thismaterial with dimethylamine in THF (5 equiv.) at room temperatureresults in the addition of the amine to the isocyanate to form the ureaat C16 and simultaneous removal of the Fmoc protecting group to provideintermediate 7-1 (R₁═R₂═CH₃). Treatment of this material withimidazole-1-sulfonylazide in the presence of potassium carbonate andcopper sulfate in methanol generates the corresponding 3′-azide.Desilylation of this material with HF/pyridine, followed by purificationby HPLC under aqueous acidic conditions produces the desired compound 7.

Specific Compounds 7 7 NR1; R2=H:

Q

R

S

T

U

Alternatively, compounds 7 can be prepared in the following manner.

Example 82. Synthesis of Compound 7-Q

A round bottom flask was charged with 2-BF (Example 4) (500 mg, 525.14mmol) which was dissolved in THF (3 mL) and MeOH (3 mL) at 20° C. K₂CO₃(290.32 mg, 2.10 mmol, 4 equiv.), CuSO₄.5H₂O (5.24 mg, 21.01 umol, 0.04equiv.) and 1H-imidazole-1-sulfonyl azide (264.18 mg, 1.26 mmol, 2.4 eq,HCl) were subsequently added and the reaction was stirred for 2 hours atRT. The reaction mixture was pour into 2-methoxy-2-methylpropane (200mL). The resulting mixture was filtered, and the filtrate wasconcentrated under reduced pressure. Purification by prep-HPLC(FA)chromatography afforded 14 mg of 7-Q as a yellow solid. ¹H NMR (400 MHz,Methanol-d4+Py-d5): δ ppm 6.09-6.57 (m, 14H), 5.49 (d, J=6.62 Hz, 2H),4.71 (s, 1H), 4.44-4.64 (m, 3H), 4.22-4.39 (m, 2H), 4.01-4.18 (m, 3H),3.51-3.91 (m, 6 H), 3.34-3.45 (m, 3H), 2.67 (s, 3H), 2.30-2.58 (m, 4H),2.14-2.28 (m, 2H), 1.94-2.09 (m, 2H), 1.25-1.93 (m, 18H), 1.21 (d,J=6.17 Hz, 3H), 1.10 (d, J=6.17 Hz, 4H), 0.99-1.05 (m, 3 H). LCMS (ESI):m/z: [M+Na] calcd for C₄₈H₇₅N₅O₁₆Na: 1000.5; found 1000.5.

Example 83. Synthesis of Compound 7-S

Compound 7-S was synthesized in the manner similar to Compound 7-Q(Example 82), except 2-BF (Example 4) was substituted with 2-AF (Example15). ¹H NMR (400 MHz, Methanol-d4+Py-d5): δ ppm 6.37-6.54 (m, 4H),6.24-6.37 (m, 5H), 6.10-6.24 (m, 4H), 5.49 (s, 1H), 5.31-5.40 (m, 2H),4.71 (s, 1H), 4.60 (m, 1H), 4.50 (m, 2H), 4.20-4.35 (m, 3H) 4.13 (d,J=3.09 Hz, 2H), 4.04 (m, 2H), 3.77-3.85 (m, 2H), 3.70 (d, J=11.03 Hz,1H), 3.55 (m, 5H), 3.41 (m, 3H), 3.22 (m, 1H), 2.51 (m, 1H), 2.32 (m,3H), 1.98 (s, 1H), 1.62-1.88 (m, 7H), 1.54 (m, 4H), 1.42 (m, 3H), 1.32(d, J=6.17 Hz, 3H), 1.21 (d, J=6.17 Hz, 3H), 1.10 (d, J=6.62 Hz, 3H),1.02 (d, J=7.06 Hz, 3H). LCMS (ESI): m/z: [M+H] calcd for C₅₃H₈₄N₆O₁₇:1077.59; found 1077.6.

Example 84. Synthesis of Compound 7-T

Compound 7-T was synthesized in the manner similar to Compound 7-Q(Example 82), except 2-BF (Example 4) was substituted with 2-AG (Example25). ¹H NMR (400 MHz, Methanol-d4+Pyr-d5): δ ppm 6.16-6.49 (m, 13H),5.53 (s, 1H), 4.74 (s., 1H), 4.26-4.60 (m., 5H), 4.17 (s., 1H),3.87-3.90 (m, 4H), 3.21-3.44 (m, 6H), 2.20-2.33 (m, 9H), 1.32-1.86 (m,15H), 1.33 (d, J=8 Hz, 3H), 1.22 (d, J=8 Hz, 3H), 1.10 (d, J=8 Hz, 3H),1.30 (d, J=8 Hz, 3H). LCMS (ESI): m/z: [M+Na] calcd for C₅₁H₇₉N₅O₁₆Na:1040.19; found 1040.4.

Example 85. C2′-epi-Urea 8

Synthesis of 8

Synthesis of 8-1

To a stirred suspension of AmB (4.0 g, 4.3 mmol, 1.0 equiv.) in DMF:MeOH(75 mL: 75 mL) in a 300 mL round bottom flask at 23° C. was addedpyridine (5.0 mL, 50.0 mmol, 11.5 equiv.) and alloc-succinimide (2.4 g,12.05 mmol, 2.8 equiv.). After stirring for 16 h at 23° C., the darkorange, homogeneous solution was slowly poured into rapidly stirringEt₂O (3.5 L). The yellow suspension was filtered through Whatman 42filter paper (110 mm diameter) and washed with Et₂O (3×100 mL) beforethe cake was allowed to fully dry. The fully dried alloc-AmB yellowpowder (4.3 mmol, quantitative) was taken on to the subsequent reactionwithout further purification.

To a stirred suspension of alloc-AmB (4.0 g, 4.3 mmol, 1.0 equiv.) inDMF:MeOH (10:1) in a 300 mL round bottom flask at 23° C. was addedsequentially Hunig's base (3.75 mL, 21.5 mmol, 5.0 equiv.) and allylbromide (11.2 mL, 129.0 mmol, 30 equiv.). After stirring for 8 h at 23°C., the dark orange, homogeneous solution was slowly poured into rapidlystirring Et₂O:Hex (1:1, 3.5 L). The subsequent yellow suspension wasfiltered through Whatman 42 filter paper (110 mm diameter) and washedwith Et₂O (3×100 mL) before the cake was allowed to fully dry. The fullydried alloc-allylester-AmB (4.3 mmol, quantitative) was taken on to thesubsequent reaction as a yellow powder without further purification.

To a stirred suspension of alloc-allylester-AmB (4.3 mmol, 1.0 equiv.)in MeOH (35 mL, 0.1 M) in a 300 mL round bottom flask at 23° C. wasadded anisaldehyde dimethylacetal (4.0 mL, 23.5 mmol, 5.5 equiv.) andstirred for 10 min until a very fine, uniform suspension formed. CSA(250 mg, 1.08 mmol, 0.25 equiv.) as a white crystalline solid was thenadded in one portion. After stirring at 23° C. for 30 min, Et₃N wasadded (˜160 μL) followed by THF (81 mL to dilute down to 0.03M). Thereaction was slowly poured into rapidly stirring hexane (3.5 L). Thesubsequent yellow suspension was filtered through Whatman 42 filterpaper (110 mm diameter) and washed with Et₂O (3×100 mL) before the cakewas allowed to fully dry. The product was purified via flashchromatography (SiO₂, gradient elution 50:49:1 EtOAc:Hex:MeOH to 75:24:1EtOAc:Hex:MeOH) to afford 8-1 (1.56 g, 1.204 mmol, 28%) as an orangesolid. R_(f)=0.21 (50:49:1) EtOAc:Hex:MeOH) Calculated for C₇₁H₉₅NO₂₁(M+Na)+: 1320.6294, Found: 1320.6285.

Synthesis of 8-2

Intermediate 8-1 (4.06 g, 3.127 mmol, 1.0 equiv.) was azeotropicallydried with benzene (3×10 mL) and placed on high vacuum overnight in a500 mL round bottom flask. To intermediate 8-1 was added THF (105 mL)followed by DIPEA (0.87 mL, 5.0 mmol, 1.6 equiv.).

In a separate 200 mL round bottom flask was added sequentially THF (64mL), DMAP (611.2 mg, 5.0 mmol, 1.6 equiv.), and dropwisep-tertbutylbenzoylchloride (855 μL, 4.38 mmol, 1.4 equiv.) forming afine, white suspension. Most of this suspension was slowly addeddropwise via cannula to the THF, DIPEA and 8-1 solution over ˜50 minuntil a majority of the starting material was converted as judged byTLC. The reaction was diluted with EtOAc and transferred to a separatoryfunnel containing aqueous saturated sodium bicarbonate and extractedwith EtOAc. The combined organic phases were dried over sodium sulfate,filtered and concentrated under reduced pressure. Purification by flashchromatography (SiO₂, gradient eluent 65:33:2 EtOAc:Hex:MeOH isocratic)afforded the desired acylated intermediate (2.28 g, 1.56 mmol, 50%yield) as an orange solid. Rf=0.24 (65:33:2 EtOAc:Hex:MeOH, HRMS (ESI)Calculated for C₈₂H₁₀₇NO₂₂ (M+Na)+: 1480.7182, Found: 1480.7172. Thisacylated intermediate (4.15 g, 2.846 mmol, 1.0 equiv.) wasazeotropically dried with benzene (3×10 mL) and placed on high vacuumovernight in a 300 mL round bottom flask. DCM (48 mL) and hexanes (48mL) were added followed by freshly distilled 2,6-lutidine (2.98 mL,25.58 mmol, 9.1 equiv.) and cooled to 0° C. Diethylisopropylsilyltriflate (DEIPSOTf; 3.39 mL, 17.05 mmol, 6.0 equiv.) was added dropwiseover 10 min and stirred for another hour at 0° C. The reaction wasdiluted with Et₂O (200 mL), transferred to a separatory funnelcontaining Et₂O and aqueous saturated bicarbonate, and extracted withEt₂O. The combined organic phases were dried over sodium sulfate,filtered, and concentrated under reduced pressure. Purification by flashchromatography (SiO₂, gradient eluent 1:9 EtOAc:Hex to 1:4 EtOAx:Hex)afforded 8-2 (4.46 g, 2.28 mmol, 80% yield) as an orange solid.R_(f)=0.21 (1:4 EtOAc:Hex) HRMS (ESI) Calculated for C₁₁₀H₁₇₁NO₂₂(M+Na)+: 1993.1268, Found: 1993.1189.

Synthesis of 8-3

Intermediate 8-2 (6.39 g, 3.24 mmol, 1.0 equiv.) was azeotropicallydried with benzene (3×10 mL) and placed on high vacuum overnight in a300 mL round bottom flask. Intermediate 8-2 was added to a mixture ofTHF (71 mL) and MeOH (140 mL). KCN (314.8 mg, 4.83 mmol, 1.5 equiv.) wasadded, and the material was placed under Ar atmosphere, sealed andwarmed to 40° C. and stirred for 48 h behind a blast shield. Thereaction transferred to a separatory funnel containing Et₂O and aqueoussaturated bicarbonate. The organic phase was washed with water followedby brine. The combined aqueous phases were extracted with Et₂O. Thecombined organic phases were dried over sodium sulfate, filtered andconcentrated under reduced pressure. Purification by flashchromatography (SiO₂, gradient eluent 1:9 EtOAc:Hex to 1:4 EtOAx:Hex)afforded the deprotected alcohol (2.93 g, 1.62 mmol, 50% yield) as anorange solid. R_(f)=0.22 (3:7 EtOAc:Hex). HRMS (ESI) Calculated forC₉₉H₁₅₉NO₂₁ (M+Na)+: 1833.0379, Found: 1833.0355. The deprotectedalcohol (2.93 g, 1.62 mmol, 1.0 equiv.) was azeotropically dried withbenzene (3×10 mL) and placed on high vacuum overnight in a 250 mL roundbottom flask. p-Nitrobenzoic acid (1.62 g, 9.7 mmol, 6.0 equiv.), PPh₃(2.54 mg, 9.7 mmol, 6.0 equiv.) and benzene (54 mL) were added. Thesolution was cooled to 0° C. and DIAD (1.91 mL, 9.7 mmol, 6.0 equiv.)was added drop-wise and the reaction was stirred at 0° C. for 1 h. Thereaction was then stirred at 23° C. for 3 h. The reaction wastransferred to a separatory funnel containing Et₂O and aqueous saturatedsodium bicarbonate. The organic phase was washed with water followed bybrine. The combined aqueous phases were extracted with Et₂O. Thecombined organic phases were dried over sodium sulfate, filtered andconcentrated under reduced pressure. Purification by flashchromatography (SiO₂, gradient eluent 1:9 EtOAc:Hex to 1:4 EtOAx:Hex)afforded the C2′epi nitrobenzoate (2.66 g, 1.36 mmol, 84% yield) as anorange solid. Rf=0.2 (1:4 EtOAc:Hex) HRMS (ESI) Calculated forC₁₀₆H₁₆₂N₂O₂₄Si₄ (M+Na)+: 1982.0492, Found: 1982.0464.

The C2′epi nitrobenzoate (2.46 g, 1.25 mmol, 1.0 equiv.) wasazeotropically dried with benzene (3×10 mL) and placed on high vacuumovernight in a 250 mL iChem. Flask THF (27.3 mL) and MeOH (54.6 mL) wereadded followed by KCN (121.8 mg, 1.87 μmol, 1.5 equiv.). The reactionwas placed under Ar atmosphere, sealed and warmed to 40° C. and stirredfor 48 h behind a blast shield. The reaction transferred to a separatoryfunnel containing Et₂O and aqueous saturated bicarbonate. The organicphase was washed with water followed by brine. The combined aqueousphases were extracted with Et₂O. The combined organic phases were driedover sodium sulfate, filtered and concentrated under reduced pressure.Purification by flash chromatography (SiO₂, gradient eluent 1:9EtOAc:Hex to 1:4 EtOAx:Hex) afforded 8-3 (1.72 g, 0.948 mmol, 76% yield)as an orange solid. Rf=0.2 (3:7 EtOAc:Hex. HRMS (ESI): Calculated forC₉₉H₁₅₉NO₂₁Si₄ (M+Na)+: 1833.0379, Found: 1833.03.

Synthesis of 8

Intermediate 8-3 is converted to the desired target 8 using standardmodifications described in Scheme 8. Specifically, simultaneous cleavageof the allyl ester and alloc groups proceeds smoothly using palladiumcatalysis and thiosalicylic acid. Reprotection of the mycosaminenitrogen and conversion of the carboxylate group to a urea providesintermediate 8-5, which is desilylated and ketalized using standardconditions to produce 8.

Example 86. In Vitro Assessment of Biological Activity

A high therapeutic index is preferable for a drug to have a favorablesafety profile. Classically, in an established clinical indicationsetting of an approved drug, therapeutic index refers to the ratio ofthe dose of drug that causes adverse effects at an incidence/severitynot compatible with the targeted indication (e.g. toxic dose in 50% ofsubjects, TD₅₀) divided by the dose that leads to the desiredpharmacological effect (e.g. efficacious dose in 50% of subjects, ED₅₀).In a drug development setting, therapeutic index is, more generally, thequantitative relationship between efficacy (pharmacology) and safety(toxicology).

Each derivative described herein is tested in vitro for biologicalactivity against both yeast and human cells to determine its therapeuticindex. A broth microdilution experiment determines the MIC (minimuminhibitory concentration) of each derivative against S. cerevisiae andthe clinically relevant C. albicans, thereby establishing the antifungalactivity of each novel derivative. To test for toxicity against humancells, each compound is tested in a hemolysis assay against red bloodcells which determines the concentration of compound required to cause90% lysis of human red blood cells (EH₉₀). Additionally, each compoundis exposed to human primary renal tubule cells to determine the toxicityof each compound against kidney cells. These assays, when comparedagainst the known or measured values of AmB against the same cells,demonstrate the improvement in therapeutic index of each compound ascompared to AmB.

Example 87. In Vivo Assessment of Biological Activity

The antifungal efficacies of compounds are tested in vivo in a mousemodel of disseminated candidiasis. In this experiment neutropenic miceare infected with C. albicans via the tail vein, and then 2 hours postinfection the mice are treated with a single intraperitoneal injectionof AmB or test agent. Then 2, 6, 12, and 24 hours post infection themice are sacrificed, and the fungal burden present in their kidneys isquantified.

INCORPORATION BY REFERENCE

All patents and published patent applications mentioned in thedescription above are incorporated by reference herein in theirentirety.

EQUIVALENTS

Having now fully described the present invention in some detail by wayof illustration and example for purposes of clarity of understanding, itwill be obvious to one of ordinary skill in the art that the same can beperformed by modifying or changing the invention within a wide andequivalent range of conditions, formulations and other parameterswithout affecting the scope of the invention or any specific embodimentthereof, and that such modifications or changes are intended to beencompassed within the scope of the appended claims.

1.-80. (canceled)
 81. A method of treating a fungal infection,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of Formula IV, thereby treating thefungal infection;

wherein, independently for each occurrence: X is —N(R²)—, —C(R³)(R³)—,or —O—; R² is hydrogen or a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; R³ ishydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or a substitutedor unsubstituted group selected from the group consisting of alkyl,cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl,heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy, amino,amido, azido, aminoalkyl, and alkoxyl; R⁵ is selected from the groupconsisting of hydrogen, alkyl, and haloalkyl; when X is —N(R²)—, R¹ is asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, acyl, amino, amido,aminoalkyl, and alkoxyl; or R¹ and R², together with the nitrogen towhich they are attached, may form a substituted or unsubstituted 3- to10-membered heterocyclic ring, wherein said ring is monocyclic,bicyclic, tricyclic, or spirocyclic; when X is —C(R³)(R³)—, R¹ ishydrogen, halogen, hydroxyl, sulfhydryl, nitro, cyano, or a substitutedor unsubstituted group selected from the group consisting of alkyl,cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl,heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy, amino,amido, azido, aminoalkyl, and alkoxyl; or the two instances of R³,together with the carbon to which they are attached, may form asubstituted or unsubstituted 3- to 10-membered aliphatic or heterocyclicring, wherein said ring is monocyclic, bicyclic, tricyclic, orspirocyclic; and when X is —O—, R¹ is a substituted or unsubstitutedgroup selected from the group consisting of alkyl, alkenyl, cycloalkyl,(cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl,aralkyl, heteroaralkyl, acyl, amino, amido, and aminoalkyl.
 82. Themethod of claim 81, wherein the compound is administered intravenously.83. The method of claim 81, wherein the compound is administered orally.84. The method of claim 81, wherein X is —N(R²)—.
 85. The method ofclaim 81, wherein X is —C(R³)(R³)—.
 86. The method of claim 81, whereinX is —O—.
 87. The method of claim 81, wherein —XR¹ is selected from thegroup consisting of

wherein, independently for each occurrence: R^(a) is hydrogen or asubstituted or unsubstituted group selected from the group consisting ofalkyl, cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl,aryl, heteroaryl, aralkyl, heteroaralkyl, acyl, amino, amido,aminoalkyl, and alkoxyl; R^(b) is hydrogen, halogen, hydroxyl,sulfhydryl, nitro, cyano, or a substituted or unsubstituted groupselected from the group consisting of alkyl, cycloalkyl,(cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl,aralkyl, heteroaralkyl, carboxyl, acyl, acyloxy, amino, amido, azido,aminoalkyl, and alkoxyl; R^(c) is hydrogen or a substituted orunsubstituted group selected from the group consisting of alkyl,cycloalkyl, (cycloalkyl)alkyl, heterocyclyl, (heterocyclyl)alkyl, aryl,heteroaryl, aralkyl, heteroaralkyl, acyl, amino, amido, and aminoalkyl;and R^(d) is hydrogen or a substituted or unsubstituted group selectedfrom the group consisting of alkyl, cycloalkyl, (cycloalkyl)alkyl,heterocyclyl, (heterocyclyl)alkyl, aryl, heteroaryl, aralkyl,heteroaralkyl, acyl, amino, amido, aminoalkyl, and alkoxyl; or, when—XR¹ is

R^(a) and R^(d), together with the nitrogen to which they are attached,may form a substituted or unsubstituted 3- to 10-membered heterocyclicring, wherein said ring is monocyclic, bicyclic, tricyclic, orspirocyclic.
 88. The method of claim 81, wherein —XR¹ is selected fromthe group consisting of


89. The method of claim 81, wherein —XR¹ is selected from the groupconsisting of


90. The method of claim 81, wherein —XR¹ is selected from the groupconsisting of


91. The method of claim 81, wherein —XR¹ is selected from the groupconsisting of methyl, ethyl, propyl, isopropyl, propenyl,


92. The method of claim 81, wherein X is —O—; and R¹ is selected fromthe group consisting of methyl, ethyl, propyl, isopropyl, propenyl,


93. The method of claim 81, wherein R⁵ is hydrogen.
 94. The method ofclaim 81, wherein R⁵ is alkyl.
 95. The method of claim 81, wherein R⁵ ishaloalkyl.
 96. The method of claim 81, wherein R² is hydrogen.
 97. Themethod of claim 81, wherein the fungal infection is caused by apathogenic yeast.
 98. The method of claim 97, wherein the pathogenicyeast is a species of a genus selected from the group consisting ofCandida and Cryptococcus.
 99. The method of claim 98, wherein thepathogenic yeast is Candida albicans, Candida tropicalis, Candidastellatoidea, Candida glabrata, Candida krusei, Candida parapsilosis,Candida guilliermondii, Candida viswanathii, or Candida lusitaniae. 100.The method of claim 98, wherein the pathogenic yeast is Cryptococcusneoformans.
 101. The method of claim 97, wherein the pathogenic yeastcauses an infection in the oral, esophageal, or vaginal mucosalmembranes.
 102. The method of claim 81, wherein the fungal infection iscaused by a non-yeast fungal pathogen.
 103. The method of claim 102,wherein the non-yeast fungal pathogen is a species of a genus selectedfrom the group consisting of Aspergillus, Rhizopus, Mucor, Histoplasma,Coccidioides, Blastomyces, Trichophyton, Microsporum, andEpidermophyton.
 104. The method of claim 103, wherein the non-yeastfungal pathogen is selected from the group consisting of Aspergillusfumigatus, Aspergillus flavus, Aspergillus niger, Histoplasmacapsulatum, Coccidioides immitis, and Blastomyces dermatitidis.